Gas exchange composite membranes and methods of use thereof

ABSTRACT

Provided herein is a gas exchange composite membrane and methods of making the same. The gas exchange composite membrane may find use in a method of exchanging gas with blood in a subject in need of blood oxygenation support, which method is also disclosed. Also provided herein are systems and kits that find use in performing the methods of exchanging gas with blood.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority benefit of U.S. Provisional ApplicationNo. 62/081,896, filed Nov. 19, 2014, which application is incorporatedherein by reference in its entirety and for all purposes.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

This invention was made with government support under Grant Nos.TR000004 and RO1 EB014315, awarded by the National Institutes of Health,and grant no. P50 FD003793, awarded by the Food and Drug Administration.The government has certain rights in the invention.

BACKGROUND

Extracorporeal membrane oxygenation (ECMO) is a respiratory supportsystem wherein blood is pumped out of the body, flowed over a gaspermeable membrane to exchange oxygen and carbon dioxide, and returnedto the systemic circulation. A typical ECMO system uses a mechanicalpump to withdraw deoxygenated blood from the patient, via large diameter(˜¼″) tubing, through a heat exchanger and oxygenator, before returningoxygen-rich blood to the body.

Unlike mechanical ventilation, which can induce barotrauma by forcingoxygen-rich air into already damaged pulmonary alveoli, ECMO directlyoxygenates blood via a synthetic membrane, allowing the lungs to heal.ECMO allows patients to rest and recover from traumatic injury,disease-induced acute respiratory distress, or prepare for lungtransplant.

SUMMARY

Provided herein is a gas exchange composite membrane and methods ofmaking the same. The gas exchange composite membrane may find use in amethod of exchanging gas with blood in a subject in need of bloodoxygenation support, which method is also disclosed. Also providedherein are systems and kits that find use in performing the methods ofexchanging gas with blood.

A gas exchange composite membrane of the present disclosure may include:i) a non-porous, gas-permeable, polymeric membrane defining a firstsurface and a second surface opposite the first surface; and ii) anon-compliant, microporous membrane defining a third surface and afourth surface opposite the third surface, wherein the microporousmembrane includes one or more gas diffusion windows, each containing anetwork of struts defining walls of a plurality of micropores, eachmicropore extending from the third surface to the fourth surface; andthe third surface is attached to the second surface, wherein the firstsurface of the polymeric membrane provides an antithrombotic surface forgas exchange over the one or more gas diffusion windows, between bloodflowing along the first surface and a gas at the second surface. In someembodiments, the first surface is substantially flat over the one ormore gas diffusion windows. In some embodiments, the composite membranehas an oxygen gas permeability against air of 5 mL STP/cmHg/m²/min ormore. In some embodiments, the composite membrane has a carbon dioxidegas permeability against air of 20 mL STP/cmHg/m²/min or more. In someembodiments, the composite membrane has an oxygen gas transfer rateagainst blood of 0.5 mL STP/cmHg/m²/min or more, at an average bloodflow speed over the first surface in the range of about 1.0 to about 10mm/sec. In some embodiments, the composite membrane has a carbon dioxidetransfer rate against blood of 2.0 mL STP/cmHg/m²/min or more, at anaverage blood flow speed over the first surface in the range of about1.0 to about 10.0 mm/sec. In some embodiments, the polymeric membranehas an average thickness in the range of 0.01 μm to 100 μm, including0.1 to 15 μm. In some embodiments, the microporous membrane has anaverage thickness in the range of 0.001 μm to 50 μm, including 0.1 to 50μm. In some embodiments, each of the one or more gas diffusion windowshas a porosity in the range of 1% to 90%, including 20% to 80%. In someembodiments, the one or more gas diffusion windows overlie an area inthe range of 1.0 mm² to 1.0 m². In some embodiments, a strut dividingadjacent micropores of the plurality of micropores have an average widthin the range of 0.005 μm to 10 μm, including 0.01 to 5.0 μm. In someembodiments, plurality of micropores is an array of micropores havingsubstantially uniform dimensions. In some embodiments, the array is aregular array of micropores. In some embodiments, a micropore of theplurality of micropores has an average width in the range of 0.005 μm to50 μm, including 0.01 to 10 μm. In some embodiments, the micropore hasan average length in the range of 0.01 μm to 100 μm, including 1.0 to100 μm. In some embodiments, the polymeric membrane is apolydimethylsiloxane (PDMS)-based polymeric membrane. In someembodiments, the first surface is functionalized with an antifoulingagent, an anticoagulant and/or an enzyme. In some embodiments, the firstsurface is functionalized with polyethylene glycol, perfluorocarbon,heparin, polysulfobetaine, or carbonic anhydrase. In some embodiments,the microporous membrane is a microporous polysilicon, silicon, siliconcarbide, or silicon nitride membrane. In some embodiments, the fourthsurface includes an anchoring strip that circumscribes each gasdiffusion window, wherein the anchoring strip protrudes out relative toareas adjacent the anchoring strip on the fourth surface. In someembodiments, the composite membrane further includes a base substrateattached to the anchoring strip.

Also provided herein is a method of making a gas exchange compositemembrane, including: a) forming a non-compliant, microporous membranedefining a first surface; b) forming a multilayered membrane-supportingstructure containing a plurality of layers superposed among each other,wherein the multilayered membrane-supporting structure defines a secondsurface containing a superficial layer of the plurality of superposedlayers, and wherein the superficial layer includes a non-porous,gas-permeable, polymeric membrane detachably disposed over an underlyinglayer; c) bonding the first surface to the second surface; and d)detaching the underlying layer from the non-porous, gas-permeable,polymeric membrane, thereby exposing a third surface of the non-porous,gas-permeable, polymeric membrane, wherein the third surface provides anantithrombotic surface for gas exchange across the composite membrane,between blood flowing along the third surface and a gas at a fourthsurface of the non-compliant, microporous membrane opposite the firstsurface. In some embodiments, the non-porous, gas-permeable, polymericmembrane is a PDMS membrane. In some embodiments, the non-compliant,microporous membrane has an average thickness in the range of 0.01 μm to100 μm, including 0.5 to 10 μm. In some embodiments, the non-porous,gas-permeable, polymeric membrane has an average thickness in the rangeof 0.001 μm to 50 μm, including 0.5 to 10 μm. In some embodiments, thenon-compliant, microporous membrane is a microporous polysilicon,silicon, silicon carbide, or silicon nitride membrane. In someembodiments, the bonding includes plasma bonding, wet chemistry, orphysical attachment of the first surface to the second surface.

In any embodiment, the forming the non-compliant, microporous membranemay include:

-   i) depositing a sacrificial layer over a fifth surface of a base    substrate, wherein the sacrificial layer is patterned to create one    or more anchor regions, each anchor region defining a window;-   ii) depositing a non-compliant membrane on the patterned sacrificial    layer; iii) etching, including dry or wet etching, the non-compliant    membrane deposited on the patterned sacrificial layer within an area    defined by the window, to form a network of struts defining walls of    a plurality of micropores in the non-compliant membrane; iv)    removing the base substrate over the area defined by the window;    and v) removing the sacrificial layer over the area defined by the    window. In some embodiments, the base substrate is a silicon or    glass base substrate. In some embodiments, the sacrificial layer is    a silicon dioxide or silicon nitride layer. In some embodiments,    depositing the sacrificial layer includes using thermal oxidation.    In some embodiments, the sacrificial layer has a thickness in the    range of 0.005 to 10 μm, including 0.1 to 10 μm. In some    embodiments, the dry etching the non-compliant membrane includes    forming a mask layer comprising a photoresist over the non-compliant    membrane, wherein the photoresist is patterned to be present over    areas of the non-compliant membrane corresponding to the network of    struts. In some embodiments, the dry etching the non-compliant    membrane includes reactive ion etching. In some embodiments, the    removing the base substrate includes using front-to-back alignment    and deep reactive ion etching. In some embodiments, the removing the    sacrificial layer includes contacting the sacrificial layer with an    acid.

In any embodiment, the forming the multilayered membrane-supportingstructure may include i) depositing a transitory polymeric membrane overa sixth surface of a support substrate, wherein the transitory polymericmembrane defines a seventh surface opposite an eighth surface andcontacting the sixth surface of the support substrate. In someembodiments, the support substrate is a support silicon substrate. Insome embodiments, the sixth surface is a surface treated to passivatethe sixth surface against adhesion to the transitory polymeric membrane,and wherein the forming the multilayered membrane-supporting structurefurther includes; ii) surface-treating the seventh surface to passivatethe seventh surface against adhesion to the non-porous, gas-permeable,polymeric membrane; and iii) depositing the non-porous, gas-permeable,polymeric membrane over the surface-treated seventh surface; and iv)detaching the support substrate from the eighth surface. In someembodiments, the transitory polymeric membrane is a transitory PDMSmembrane. In some embodiments, the sixth surface is silanized. In someembodiments, the surface treating the seventh surface includessilanizing the seventh surface. In some embodiments, the detaching theunderlying layer from the polymeric membrane includes mechanicallydetaching the transitory polymeric membrane from the non-porous,gas-permeable, polymeric membrane. In some embodiments, the surfacetreating the seventh surface includes depositing a water-soluble polymerlayer over the seventh surface. In some embodiments, the detaching theunderlying layer from the polymeric membrane includes contacting thewater-soluble polymer layer with water to dissolve the water-solublepolymer layer. In some embodiments, the transitory polymeric membrane isa dissolvable polymeric membrane, and wherein the detaching theunderlying layer from the non-porous, gas-permeable, polymeric membraneincludes contacting the dissolvable polymeric membrane with a solvent,thereby dissolving the dissolvable polymeric membrane. In someembodiments, the dissolvable polymeric membrane is an epoxy-baseddissolvable polymeric membrane. In some embodiments, the dissolvablepolymeric membrane is a photoresist membrane. In some embodiments, thedissolvable polymeric membrane is an SU-8 membrane. In some embodiments,the solvent is an organic solvent. In some embodiments, the solvent isacetone.

In any embodiment the third surface may be a functionalized surface.

In any embodiment the method may further include functionalizing asurface of the non-porous, gas-permeable, polymeric membrane.

Also provided herein is a blood oxygenation device that includes: 1) ablood channel comprising a first inlet at a first end and a first outletat a second end opposite the first end, wherein the blood channel isconfigured to pass a flow of blood from the first inlet to the firstoutlet; 2) a first gas channel configured to pass a flow of gas; and 3)a first gas exchange composite membrane of the present disclosure,wherein the first composite membrane is disposed between the bloodchannel and the first gas channel in a manner sufficient to provide agas permeable barrier between the blood channel and the first gaschannel. In some embodiments, the device further includes: 4) a secondgas channel comprising a second inlet at a fifth end and a second outletat a sixth end opposite the fifth end, wherein the second gas channel isconfigured to pass a flow of the gas from the second inlet to the secondoutlet; and 5) a second gas exchange composite membrane, wherein thesecond composite membrane is disposed between the blood channel and thesecond gas channel in a manner sufficient to provide a gas permeablebarrier between the blood channel and the second gas channel. In someembodiments, the blood channel has a length defined by the distancebetween the first end and a second end in the range of 0.1 to 300 mm,including 1.0 to 300 mm. In some embodiments, a cross-section in a planeperpendicular to the average direction of flow of the blood in the bloodchannel is a rectangular cross-section defining a width and a height ofthe blood channel, wherein an edge of the rectangular cross-sectiondefining the width includes the first surface of the non-porous,gas-permeable, polymeric membrane of the first composite membrane. Insome embodiments, the width of the blood channel is in the range of 0.05to 300 mm, including 0.5 to 300 mm. In some embodiments, the height ofthe blood channel is in the range of 0.001 to 300 mm, including 0.01 to2.0 mm. In some embodiments, the ratio of the width to height of theblood channel is in the range of 10 to 1,000. In some embodiments, thegas diffusion windows of the first composite membrane collectivelyoverlie an area in the range of 1.0 mm² to 0.5 m². In some embodiments,the blood channel has a volume in the range of 1.0 mm³ to 1.5 m³. Insome embodiments, the flow of blood has an average direction that issubstantially perpendicular to a direction of the flow of gas. In someembodiments, the flow of blood has an average direction that issubstantially parallel to a direction of the flow of gas. In someembodiments, the device is stackable. In some embodiments, the bloodchannel includes a tapered inlet. In some embodiments, the blood channelincludes a tapered outlet. In some embodiments, the blood channelincludes a polymeric or metal channel. In some embodiments, the bloodchannel includes a polycarbonate, polyurethane or silicone channel. Insome embodiments, the blood channel is a PDMS channel. In someembodiments, the blood channel is a titanium alloy channel. In someembodiments, the channel includes a surface that is functionalized. Insome embodiments, the surface is functionalized with polyethyleneglycol, perfluorocarbon and/or heparin.

Also provided herein is a method of exchanging gas with blood includinga first dissolved gaseous compound, the method including: (A) pumpingblood from a circulatory system of a subject to an extracorporeal bloodcircuit to generate a circulating flow of the blood, the extracorporealblood circuit comprising one or more non-circuitous blood channels ofone or more blood oxygenating devices, each blood channel defining afirst end and a second end opposite the first end, wherein each of theblood oxygenating devices includes: a gas channel configured to pass aflow of gas, wherein the gas includes a second gaseous compound; one ormore gas exchange composite membranes configured to exchange gaseouscompounds between the blood and the gas across a planar surfaceseparating the blood channel and the gas channel; and (B) flowing thegas through the gas channel, thereby exchanging gaseous compoundsbetween the circulating flow of the blood and the gas. In someembodiments, a cross-section in a plane perpendicular to the averagedirection of the circulating flow of the blood in the blood channel is arectangular cross-section defining a width and a height of the bloodchannel, wherein an edge of the rectangular cross-section defining thewidth includes the planar surface separating the blood channel and thegas channel. In some embodiments, the width of the blood channel is inthe range of 0.001 to 300 mm, including 0.5 to 300 mm. In someembodiments, the height of the blood channel is in the range of 0.001 to300 mm, including 0.01 to 2.0 mm. In some embodiments, the ratio of thewidth to height of the blood channel is in the range of 10 to 1,000. Insome embodiments, the partial pressure of the second gaseous compound inthe gas is 20 cmHg or more. In some embodiments, blood is pumped at aflow rate in the range of 0.1 to 100 ml/min. In some embodiments, theflow of blood within the blood channel has a maximum shear stress of1,000 dyne cm⁻² or less. In some embodiments, the flow of blood acrosseach of the non-circuitous blood channels has a hydraulic pressure dropbetween the first end and the second end of 100 mmHg or less. In someembodiments, the one or more blood oxygenating devices has an oxygentransfer rate between the gas and the blood of 0.5 mL STP/cmHg/m²/min ormore, at an average blood flow rate in the rage of about 0.1 to 1.0mL/min. In some embodiments, the one or more blood oxygenating deviceshas an carbon dioxide transfer rate between the gas and the blood of 2.0mL STP/cmHg/m²/min or more, at an average blood flow speed over thefirst surface in the range of about 0.1 to 1.0 mL/min. In someembodiments, the one or more blood oxygenating devices collectively havea gas exchange surface area in the range of 0.01 to 10 m², including 0.1to 5 m². In some embodiments, the one or more blood oxygenating devicesincludes one or more gas exchange composite membranes.

Provided herein is a system for exchanging gas with blood using one ormore blood oxygenation devices of the present disclosure. In someembodiments, the one or more blood oxygenating devices collectivelyprovide a gas exchange surface area in the range of 0.1 to 5 m². In somecases, the one or more blood oxygenation devices are configured to bewearable.

Also provided is a kit that includes a gas exchange composite membraneof the present disclosure.

BRIEF DESCRIPTION OF THE FIGURES

The skilled artisan will understand that the drawings, described below,are for illustration purposes only. The drawings are not intended tolimit the scope of the present teachings in any way.

FIG. 1 is a schematic drawing showing a blood oxygenation device thatincludes a gas exchange composite membrane, according to embodiments ofthe present disclosure.

FIG. 2 is a schematic diagram showing a process of making anon-compliant, microporous membrane for use in a gas exchange compositemembrane, according to embodiments of the present disclosure.

FIGS. 3A-3C are schematic diagrams showing processes of making a gasexchange composite membrane, according to embodiments of the presentdisclosure.

FIGS. 4A-4D are images showing different views of a gas exchangecomposite membrane, according to embodiments of the present disclosure.

FIG. 5 is a schematic diagram showing a configuration of blood and gasflowing over a gas exchange composite membrane, according to embodimentsof the present disclosure.

FIGS. 6A and 6B are a collection of images showing a blood oxygenationdevice that includes a gas exchange composite membrane, according toembodiments of the present disclosure.

FIGS. 7A and 7B are schematic diagrams showing systems for oxygenatingblood using a blood oxygenation device that includes a gas exchangecomposite membrane, according to embodiments of the present disclosure.

FIG. 8 is a graph showing gas permeability of a gas exchange compositemembrane, according to embodiments of the present disclosure.

FIG. 9 is a graph showing pressure drop between an inlet and an outletof blood flowing through a blood oxygenation device that includes a gasexchange composite membrane in ex vivo tests, according to embodimentsof the present disclosure.

FIGS. 10A and 10B are a collection of graphs showing oxygen partialpressure and oxygen exchange rates, respectively, in blood flowingthrough a blood oxygenation device that includes a gas exchangecomposite membrane in ex vivo tests, according to embodiments of thepresent disclosure.

FIGS. 11A and 11B are a collection of graphs showing carbon dioxidepartial pressure and carbon dioxide exchange rates, respectively, inblood flowing through a blood oxygenation device that includes a gasexchange composite membrane in ex vivo tests, according to embodimentsof the present disclosure.

FIG. 12 is a graph showing pressure drop between an inlet and an outletof blood flowing through a blood oxygenation device that includes a gasexchange composite membrane in in vivo tests, according to embodimentsof the present disclosure.

FIGS. 13A and 13B are a collection of graphs showing oxygen partialpressure and oxygen exchange rates, respectively, in blood flowingthrough a blood oxygenation device that includes a gas exchangecomposite membrane in in vivo tests, according to embodiments of thepresent disclosure.

FIGS. 14A and 14B are a collection of graphs showing carbon dioxidepartial pressure and carbon dioxide exchange rates, respectively, inblood flowing through a blood oxygenation device that includes a gasexchange composite membrane in in vivo tests, according to embodimentsof the present disclosure.

FIGS. 15A and 15B are a collection of images showing a surface profileof a gas exchange composite membrane after flowing blood through a bloodoxygenation device that includes the gas exchange composite membrane.

FIG. 16 is a graph showing gas permeability of a gas exchange compositemembrane, according to embodiments of the present disclosure.

FIG. 17 is a graph showing oxygen transfer rate of a gas exchangecomposite membrane, according to embodiments of the present disclosure.

FIG. 18 is a graph showing blood oxygen saturation over time in a closedloop system when exposed to a gas exchange composite membrane, accordingto embodiments of the present disclosure.

FIG. 19 is a graph showing oxygen permeability of a gas exchangecomposite membrane in ex vivo tests, according to embodiments of thepresent disclosure.

FIG. 20 is a diagram showing a blood oxygenating device, according toembodiments of the present disclosure.

DEFINITIONS

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this disclosure belongs. Although any methods andmaterials similar or equivalent to those described herein can also beused in the practice or testing of the present teachings, some exemplarymethods and materials are now described.

A “plurality” contains at least 2 members. In certain cases, a pluralitymay have at least 10, at least 100, at least 1000, at least 10,000, atleast 100,000, at least 10⁶, at least 10⁷, at least 10⁸ or at least 10⁹or more members.

“Substantially” as used herein, may be applied to modify anyquantitative representation that could permissibly vary withoutresulting in a change in the basic function to which it is related. Forexample, a direction of flow of gas may be somewhat off-perpendicularfrom the direction of flow of blood if the gas exchange propertiesbetween the gas and the blood are not materially altered.

“Blood oxygenation” as used herein, may refer to a general process ofexchanging gas with blood of a subject to achieve gas exchange thattypically occurs in the lung. Blood oxygenation may include increasingthe oxygen content of blood and/or reduce the carbon dioxide content ofblood.

“Non-porous” as used herein, may be applied to describe a property of astructure or material that is not permeable to liquids under normalconditions for use intended in the present disclosure.

“Polymeric” as used herein, may be used to describe an organic compoundcomposed of repeating units of one or more monomers containing carbonand hydrogen atoms The monomers can also include other atoms such as Si,O, N, P, and S. A polymer may have a solid bulk polymer matrix.

“Membrane” and “film” are used interchangeably to refer to a solidmaterial that, when laid out on a planar surface, can have asubstantially planar geometry. The membrane may have one dimension (the“depth” or “thickness”) that is considerably shorter than the other twodimensions (the “width” and “length”), so as to form the planargeometry. The “surface” of a membrane refers to the area of the membranedefined by edges along the width and the length.

“Non-compliant” as used herein, may be applied to describe the propertyof a structure or material that does not substantially deform underforce experienced by the structure or material under normal conditionsfor use intended in the present disclosure. A non-compliant material mayhave a Young's modulus (E) of 100 GPa or more, e.g., 120 GPa or more,140 GPa or more, including 160 GPa or more.

“Microporous” may be used to describe a pore whose opening size has alateral dimension (i.e., the diameter, width, or length) that is at amicrometer scale (i.e., between 1.0 to 1,000 μm).

A “network” as used in reference to struts defining micropores, maydescribe struts that are interconnected.

An “array”, as used herein, refers to an arrangement of elements wherethe location of each element is spatially defined (i.e, not random). A“regular array” refers to an array that contains a uniformly repeatedarrangement of elements.

“Superficial” as used herein, may be applied to describe a layer withina multi-layered structure that is either the first or the last layer.Thus, a superficial layer of a multi-layered structure is contacted withan underlying layer on only one surface.

“Superposed” as used herein, may be used to describe a relative positionbetween at least two structures where a first surface of a firststructure contacts a second surface of a second structure, andsubstantially covers the second surface.

“Extracorporeal”, as used herein, may be applied to describe aphysiological process of a body that is replaced or supplemented by anartificial system that can perform at least some aspects of thephysiological process. For a continuous physiological process, such ascirculation of blood, the artificial system can be configured tocontinuously perform the physiological process by having physical accessto the body part relevant for the physiological process.

“Circulating”, as used herein, may be used to describe a unidirectionalmovement of a material through a closed system, where material startingat a location in the closed system returns to the same location aftermoving through the closed system.

“Non-circuitous” as used herein, may be applied to describe a shape orpath through a structure that does not meander or turn significantly.Thus, in some cases, “non-circuitous” may be interchangeable with“substantially straight.”

Before the various embodiments are described, it is to be understoodthat the teachings of this disclosure are not limited to the particularembodiments described, and as such can, of course, vary. It is also tobe understood that the terminology used herein is for the purpose ofdescribing particular embodiments only, and is not intended to belimiting, since the scope of the present teachings will be limited onlyby the appended claims.

The section headings used herein are for organizational purposes onlyand are not to be construed as limiting the subject matter described inany way. While the present teachings are described in conjunction withvarious embodiments, it is not intended that the present teachings belimited to such embodiments. On the contrary, the present teachingsencompass various alternatives, modifications, and equivalents, as willbe appreciated by those of skill in the art.

Where a range of values is provided, it is understood that eachintervening value, to the tenth of the unit of the lower limit unlessthe context clearly dictates otherwise, between the upper and lowerlimit of that range and any other stated or intervening value in thatstated range is encompassed within the present disclosure.

The citation of any publication is for its disclosure prior to thefiling date and should not be construed as an admission that the presentclaims are not entitled to antedate such publication by virtue of priorinvention. Further, the dates of publication provided can be differentfrom the actual publication dates which can need to be independentlyconfirmed.

It must be noted that as used herein and in the appended claims, thesingular forms “a,” “an,” and “the” include plural referents unless thecontext clearly dictates otherwise. It is further noted that the claimscan be drafted to exclude any optional element. As such, this statementis intended to serve as antecedent basis for use of such exclusiveterminology as “solely,” “only” and the like in connection with therecitation of claim elements, or use of a “negative” limitation.

As will be apparent to those of skill in the art upon reading thisdisclosure, each of the individual embodiments described and illustratedherein has discrete components and features which can be readilyseparated from or combined with the features of any of the other severalembodiments without departing from the scope or spirit of the presentteachings. Any recited method can be carried out in the order of eventsrecited or in any other order which is logically possible.

One with skill in the art will appreciate that the present invention isnot limited in its application to the details of construction, thearrangements of components, category selections, weightings,pre-determined signal limits, or the steps set forth in the descriptionor drawings herein. The invention is capable of other embodiments and ofbeing practiced or being carried out in many different ways.

DETAILED DESCRIPTION

As summarized above, the present disclosure provides a gas exchangecomposite membrane that may be used to oxygenate blood. The present gasexchange composite membrane may find use in providing a compact,wearable or portable system for extracorporeal membrane oxygenation, forpatients in need of blood oxygenation and carbon dioxide removal duringacute respiratory distress (FIG. 1).

The gas exchange composite membrane may include a non-porous,gas-permeable, polymeric membrane attached to a surface of anon-compliant, microporous membrane. The microporous membrane mayinclude micron-sized pores that extend through the microporous membraneand are defined by struts that form the walls of the micropores. Thus,the attachment between the polymeric membrane and the microporousmembrane includes attachment between the polymeric membrane and thestruts of the microporous membrane.

Blood may flow over the surface of the composite membrane on thenon-porous, gas-permeable, polymeric membrane side, and gaseouscompounds may be exchanged between the blood flow and a gas on the otherside of the composite membrane that has the non-compliant, microporousmembrane (FIG. 5). Because the non-porous, gas-permeable, polymericmembrane is attached to the non-compliant, microporous membrane, thelatter serves as a structural support to significantly reduce gaspressure-induced deformation of the polymeric membrane that would createobstructions to the blood flow over the polymeric membrane, and preventcollapse of the blood flow compartment due to the gas pressure. Themicropores define areas through which gas can diffuse, across thecomposite membrane, and thus provide a sufficient diffusion surface areato oxygenate blood. At the same time, the polymeric membrane prevents orreduces the risk of gas embolism in the flowing blood even at highpressure gradients of gas across the composite membrane. The polymericmembrane also prevents blood components (e.g., cells, proteins, plasma,etc) from entering the gas flow compartment.

Further aspects of the present composite membrane are now described.

Composite Membranes

The gas exchange composite membrane of the present disclosure may bedescribed with reference to the accompanying figures. With reference toFIG. 3A, a schematic diagram of a gas exchange composite membrane 300 isshown. The gas exchange composite membrane may include a non-porous,gas-permeable, polymeric membrane 330 attached to a surface of anon-compliant, microporous membrane 220. The polymeric membrane 330 maybe non-porous, such that the liquids, such as blood cannot pass throughthe polymeric membrane. The polymeric membrane may include a suitablematerial and have a suitable thickness (“h”) to allow sufficient gaspermeability, e.g., oxygen and/or carbon dioxide permeability, acrossthe composite membrane, between blood on the polymeric membrane side andgas on the microporous membrane 220 side. The polymeric membrane can bemade of any suitable material to allow gas diffusion, and to have acontrollable thickness during manufacture.

The top surface of the non-porous, gas-permeable, polymeric membrane 330may be a substantially flat surface. The substantially flat surface ofthe polymeric membrane can provide a controlled surface over which bloodflows, and allow control of blood shear, which is a factor in thrombusformation and hemolysis. Thus, the planar surface of the gas exchangecomposite membrane 300 over which blood flows, i.e., the surface of thenon-porous, gas-permeable, polymeric membrane opposite the surfaceattached to the non-compliant, microporous membrane 220, can provide asurface with reduced risk of thrombus formation and/or hemolysis than apolymeric membrane surface that is not supported by a microporousmembrane.

The microporous membrane 220 has dimensions, e.g., thickness, and ismade of material sufficient to render the microporous membranenon-compliant in response to force that the microporous membrane mayexperience during normal use of the composite membrane for gas exchangewith blood. The non-compliant microporous membrane can have a stiffnessthat resists deformation when pressure is applied to the polymericmembrane 330 attached to the non-compliant microporous membrane. Thus,in some cases, the microporous membrane is a substantially rigidmembrane. The thickness (“b”) of the microporous membrane is a suitablethickness to provide sufficient stiffness to the microporous membrane.The microporous membrane can be made of any suitable material to providethe structural support to the polymeric membrane and to havecontrollable thickness and pore sizes during manufacture.

With reference to FIG. 2, (4), the non-compliant, microporous membrane220 may include a number of struts 222 that define walls of a number ofmicropores 223. The struts may be interconnected to form a network ofstruts (see also, FIG. 4B). As such, the shape of the struts and theirinterconnectivity can define the shape and the pattern of themicropores. The micropores each extend from one surface of themicroporous membrane to the other surface. The path through the centerof a pore may follow any suitable path, and may be substantiallyperpendicular to the plane of the surfaces (see also, FIG. 4C). Gasexchange can occur across the area of the composite membrane 300 thatcorresponds to the area where the struts define the micropores in thenon-compliant, microporous membrane 220, which area can be defined as agas diffusion window. The non-compliant, microporous membrane may beattached to a base substrate 240 in a manner such that the basesubstrate does not occlude the gas diffusion window.

The polymeric membrane 330 can have any suitable thickness (FIG. 3A,“h”). In some cases, the average thickness of the polymeric membrane is0.1 μm or more, e.g., 0.5 μm or more, 1.0 μm or more, 2.0 μm or more,3.0 μm or more, including 4.0 μm or more, and is 15 μm or less, e.g., 10μm or less, 8.0 μm or less, 6.0 μm or less, including 5.0 μm or less. Incertain embodiments, the average thickness of the polymeric membrane isin the range of 0.1 to 25 μm, e.g., 0.5 to 15 μm, 1.0 to 10 μm, 2.0 to8.0 μm, including 3.0 to 6.0 μm.

The microporous membrane 220 can have any suitable thickness (FIG. 2,“b”). In some cases, the average thickness of the microporous membraneis 0.1 μm or more, e.g., 0.2 μm or more, 0.5 μm or more, 1.0 μm or more,including 5.0 μm or more, and is 50 μm or less, e.g., 20 μm or less, 10μm or less, 5.0 μm or less, including 4.0 μm or less. In certainembodiments, the average thickness of the microporous membrane is in therange of 0.1 to 100 μm, e.g., 0.2 to 50 μm, 0.2 to 20 μm, 0.2 to 10 μm,including 0.5 to 5.0 μm.

The gas diffusion window of the microporous membrane 220 can have anysuitable porosity. In some cases, the gas diffusion window has aporosity of 20% or more, e.g., 30% or more, 40% or more, 50% or more,including 60% or more, and is 80% or less, e.g., 70% or less, 60% orless, including 50% or less. In some cases, the gas diffusion window hasa porosity in the range of 20 to 80%, e.g., 30 to 70%, including 30 to60%.

The gas diffusion window of the microporous membrane 220 can overlie anysuitably-sized area of the present composite membrane 300. In somecases, the gas diffusion window overlies an area of 1.0 mm² or more,e.g., 10 mm² or more, 100 mm² or more, 1,000 mm² or more, including 0.01m² or more, and overlies an area of 1.0 m² or less, e.g., 0.1 m² orless, 0.01 m² or less, including 1,000 mm². In some cases, the gasdiffusion window overlies an area in the range of 1.0 mm² to 1.0 m²,e.g., 10 mm² to 0.1 m², 100 mm² to 0.1 m², including 1,000 mm² to 0.1m².

The width (FIG. 2, “d”) of a strut dividing adjacent micropores can haveany width. In certain cases, the average width of a strut dividingadjacent micropores is 0.01 μm or more, e.g., 0.1 μm or more, 0.2 μm ormore, 0.3 μm or more, including 0.4 μm or more, and is 5.0 μm or less,4.0 μm or less, 3.0 μm or less, 2.0 μm or less, including 1.0 μm orless. In certain cases, the average width of a strut dividing adjacentmicropores is in the range of 0.01 to 5.0 μm, e.g., 0.1 to 4.0 μm, 0.2to 3.0 μm, including 0.2 to 2.0 μm.

The micropores 223 may be arranged across the surface of the microporousmembrane in any suitable manner. In some cases, the micropores are anarray of micropores. In some cases, the micropores have a uniform widthand lengths across the microporous membrane. In certain cases, themicropores are in a regular array.

The pore shape of the micropores 223 may have any suitable shape anddimensions. The pores shape of the micropores may be substantiallycircular, oval, rectangular, square, triangular, etc. A micropore mayhave an average width (FIG. 2, “e”) of 0.01 μm or more, e.g., 0.1 μm ormore, 0.2 μm or more, 0.3 μm or more, 0.4 μm or more, including 0.5 μm,10.0 μm or more, and may have an average width of 10 μm or less, e.g.,5.0 μm or less, 3.0 μm or less, 2.0 μm or less, 1.0 μm or less,including 0.8 μm or less. In certain embodiments, a micropore may havean average width in the range of 0.01 to 10 μm, e.g., 0.1 to 10 μm, 0.2to 5.0 μm, 0.2 to 3.0 μm, 0.3 to 1.0 μm, including 0.3 to 0.8 μm. Amicropore may have an average length of 1.0 μm or more, e.g., 2.0 μm ormore, 3.0 μm or more, 4.0 μm or more, including 5.0 μm or more, and mayhave an average width of 100 μm or less, e.g., 50 μm or less, 30 μm orless, 20 μm or less, 10 μm or less, including 8.0 μm or less. In certainembodiments, a micropore may have an average width in the range of 1.0to 100 μm, e.g., 2.0 to 50 μm, 2.0 to 30 μm, 3.0 to 20 μm, 3.0 to 10 μm,including 3.0 to 8.0 μm.

The polymeric membrane 330 may be any suitable polymeric material foruse in the present composite membrane as a non-porous, gas-permeablepolymeric membrane. The polymeric membrane can be a biocompatiblepolymeric membrane. Suitable material for a non-porous, gas-permeablepolymeric membrane include, but are not limited to polypropylene,polymethylpentene, polyisoprene, polybutadiene, polychloroprene,polyisobutylene, poly(styrene-butadiene-styrene), polyurethane, silicon,poly(bis(fluoroalkoxy)phosphazene), poly(carboranesiloxanes),poly(acrylonitrile-butadiene), poly(l-butene),poly(chlorotrifluoroethylene-vinylidene fluoride) copolymers, poly(ethylvinyl ether), poly(vinylidene fluoride), poly(vinylidenefluoride-hexafluoropropylene) copolymer, polyvinylchloride (PVC),polysulfone, polycarbonate, polymethylmethacrylate (PMMA), orpolytetrafluoroethylene.

In some cases, the polymeric membrane 330 is a silicone membrane. Insome cases, the polymeric membrane is a vulcanizing silicone, e.g., aroom temperature vulcanizing (RTV) silicone. In some cases, thepolymeric membrane is polydimethylsiloxane (PDMS), and derivativesthereof. In some cases, the polymeric membrane may include any solidPDMS polymer composed of at least one dimethylsiloxane monomer. In someinstances, at least two dimethylsiloxane monomers are used to make thePDMS substrate. The monomer mixtures may contain additional components,such as other monomers or a catalyst, such as platinum. Various monomermixtures are commercially available and include, for example, Sylgard®184 (Dow Corning Corporation, Midland, Mich., United States), RTV 615(Sil-Mid limited, Coleshill, West Midlands, United Kingdom) andELASTOSiL® RT 601 (Wacker Chemie AG, San Jose, Calif., United States).The ratio of monomer to crosslinker used to make the polymer membranecontaining PDMS may be any suitable ratio. In some cases, the ratio ofmonomer to crosslinker is 1 or more, e.g., 2 or more, 3 or more, 4 ormore, 5 or more, including 8 or more, and is 20 or less, e.g., 15 orless, 13 or less, 12 or less, 11 or less, including 10 or less. In somecases, the ratio of monomer to crosslinker is in the range of 1 to 20,e.g. 1 to 15, 2 to 13, including 2 to 12. The PDMS may be cured at anysuitable temperature. In some cases, the PDMS is cured at 60° C. ormore, e.g., 70° C. or more, 75° C. or more, including 80° C. or more,and is cured at 90° C. or less, e.g., 85° C. or less, including 80° C.or less. In some cases, the PDMS is cured at a temperature in the rangeof 60 to 90° C., e.g., 70 to 90° C., including 75 to 85° C. The PDMS maybe cured for any suitable amount of time. In some cases, the PDMS iscured for 30 min or more, e.g., 1.0 hr or more, 1.5 hrs or more,including 2.0 hrs or more, and is cured for 24 hrs or less, e.g., 12 hrsor less, 8.0 hrs or less, 4.0 hrs or less, including 2.0 hrs or less. Insome cases, the PDMS is cured for 0.5 to 24 hrs, e.g., 1.0 to 12 hrs,1.0 to 8.0 hrs, including 1.0 to 4.0 hrs.

The surface of the polymeric membrane 330, e.g., the surface oppositethe surface attached to the microporous membrane 220, may be a surfacefunctionalized to reduce fouling of the membrane surface, reducecoagulation of blood on the surface of the membrane surface, and/orincrease the efficiency of gas exchange of a gaseous compound in theblood, such as carbon dioxide. In some cases, the surface of thepolymeric membrane is functionalized with polyethylene glycol,perfluorocarbon, heparin, and/or carbonic anhydrase. Any other suitablefunctionality may be provided to the surface of the polymeric membrane.

The microporous membrane 220 may include any suitable material for useas a non-compliant microporous membrane. Suitable materials include, butare not limited to, ceramics, glass, glass polymers, glass/polymermaterials, metals (e.g., chromium, cobalt, gold, molybdenum, nickel,stainless steel, titanium, tungsten steel, and the like), moldedplastics, polysilicon, silicon-based organic polymers.

In some cases, the composite membrane 300 includes an anchoring strip221 that protrudes from the surface of the microporous membrane 220opposite the surface attached to the polymeric membrane 330. Theanchoring strip may protrude out relative to areas adjacent theanchoring strip on the surface of the microporous membrane by anysuitable distance, and may protrude out by 0.1 μm or more, e.g., 0.2 μmor more, 0.5 μm or more, 0.75 μm or more, including 1.0 μm or more, andmay protrude out by 5.0 μm or less, e.g., 4.0 μm or less, 3.0 μm orless, including 2.0 μm or less. In some cases, the anchoring strip mayprotrude out by a range of 0.1 to 5.0 μm, e.g., 0.2 to 4.0 μm 0.5 to 3.0μm, including 0.75 to 2.0 μm. The anchoring strip may circumscribe a gasdiffusion window of the composite membrane. In some cases, the compositemembrane includes a base substrate 240 attached to the anchoring strip.The base substrate is any suitable base substrate on which to constructthe microporous membrane. In some cases, the base substrate is a siliconwafer.

The base substrate 240 may have any suitable thickness (“c” in FIG. 2).In some embodiments, the base substrate has a thickness of 10 μm ormore, e.g., 50 μm or more, 100 μm or more, 200 μm or more, 300 μm ormore, including 400 μm or more, and has a thickness of 1,000 μm or less,e.g., 800 μm or less, 600 μm or less, including 500 μm or less. In somecases, the base substrate has a thickness in the range of 10 to 1,000μm, e.g., 50 to 800 μm, 100 to 600 μm, including 200 to 600 μm.

The composite membrane can have any suitable permeability to gaseouscompounds that are to be exchanged with, e.g., blood. Gas permeabilitymay be measured using a dry flow cell connected to a pressurized gassupply containing the gaseous compound of interest, and a bubble flowmeter. In some embodiments, the present composite membrane has an oxygengas permeability against air of 5 mL Standard temperature and pressure(STP)/cmHg/m²/min or more, e.g., 7 mL STP/cmHg/m²/min or more, 9 mLSTP/cmHg/m²/min or more, 10 mL STP/cmHg/m²/min or more, including 11 mLSTP/cmHg/m²/min or more, and has an oxygen gas permeability against airof 50 mL STP/cmHg/m²/min or less, e.g., 40 mL STP/cmHg/m²/min or less,30 mL STP/cmHg/m²/min or less including 20 mL STP/cmHg/m²/min or less.In some embodiments, the present composite membrane has an oxygen gaspermeability against air in the range of 5 to 50 STP/cmHg/m²/min, e.g.,5 to 40 mL STP/cmHg/m²/min, 5 to 30 mL STP/cmHg/m²/min, including 10 to20 mL STP/cmHg/m²/min.

In some embodiments, the present composite membrane has a carbon dioxidegas permeability against air of 20 mL STP/cmHg/m²/min or more, e.g., 40mL STP/cmHg/m²/min or more, 50 mL STP/cmHg/m²/min or more, 55 mLSTP/cmHg/m²/min or more, including 60 mL STP/cmHg/m²/min or more, andhas a carbon dioxide gas permeability against air of 200 mLSTP/cmHg/m²/min or less, e.g., 150 mL STP/cmHg/m²/min or less, 100 mLSTP/cmHg/m²/min or less including 90 mL STP/cmHg/m²/min or less. In someembodiments, the present composite membrane has a carbon dioxide gaspermeability against air in the range of 20 to 200 STP/cmHg/m²/min,e.g., 40 to 150 mL STP/cmHg/m²/min, 50 to 100 mL STP/cmHg/m²/min,including 60 to 90 mL STP/cmHg/m²/min.

The present composite membrane has any suitable oxygen gas transfer rateagainst blood at an average blood flow speed over the first surface inthe range of about 1.0 to about 10 mm/sec. In some embodiments, thecomposite membrane has an oxygen gas transfer rate against blood of 0.5mL STP/cmHg/m²/min or more, e.g., 1.0 mL STP/cmHg/m²/min or more, 1.5 mLSTP/cmHg/m²/min or more, including 2.0 mL STP/cmHg/m²/min or more, andhas an oxygen gas transfer rate against blood of 10 mL STP/cmHg/m²/minor less, e.g., 8.0 mL STP/cmHg/m²/min or less, 6.0 mL STP/cmHg/m²/min orless, 4.0 mL STP/cmHg/m²/min or less, including 3.0 mL STP/cmHg/m²/minor less. In some embodiments, the composite membrane has an oxygen gastransfer rate against blood in the range of 0.5 to 10 mLSTP/cmHg/m²/min, e.g., 1.0 to 8.0 mL STP/cmHg/m²/min, 1.5 to 6.0 mLSTP/cmHg/m²/min, including 1.5 to 4.0 mL STP/cmHg/m²/min.

In some embodiments, the composite membrane has a carbon dioxide gastransfer rate against blood of 2.0 mL STP/cmHg/m²/min or more, e.g., 2.5mL STP/cmHg/m²/min or more, 3.0 mL STP/cmHg/m²/min or more, 4.0 mLSTP/cmHg/m²/min or more, including 5.0 mL STP/cmHg/m²/min or more, andhas a carbon dioxide gas transfer rate against blood of 50 mLSTP/cmHg/m²/min or less, e.g., 40 mL STP/cmHg/m²/min or less, 30 mLSTP/cmHg/m²/min or less, 20 mL STP/cmHg/m²/min or less, including 10 mLSTP/cmHg/m²/min or less. In some embodiments, the composite membrane hasa carbon dioxide transfer rate against blood in the range of 2.0 to 50mL STP/cmHg/m²/min, e.g., 3.0 to 30 mL STP/cmHg/m²/min, 4.0 to 20 mLSTP/cmHg/m²/min, including 5.0 to 10 mL STP/cmHg/m²/min.

Methods of making a Composite Membrane

In general terms, the fabrication of a gas exchange composite membraneof the present disclosure includes a) forming a non-compliant,microporous membrane defining a first surface; b) forming a multilayeredmembrane-supporting structure having a plurality of superposed layers,wherein the multilayered membrane supporting structure defines a secondsurface of a superficial layer of the plurality of superposed layers,wherein the superficial layer includes a non-porous, gas-permeable,polymeric membrane detachably disposed over an underlying layer; c)bonding the first surface to the second surface; and d) detaching thepolymeric membrane from the underlying layer. Various embodiments ofmaking a gas exchange composite membrane of the present disclosure aredescribed with reference to the accompanying drawings.

With reference to FIG. 2, a method of fabricating a non-compliant,microporous membrane 220 of the present composite membrane is described.The method may include constructing a multilayered structure on thefront side of a base substrate 240, e.g., a silicon wafer, where asacrificial layer 230, e.g., a silicon dioxide layer, is deposited overthe front side of the base substrate, and a non-compliant membrane 220,e.g., a polysilicon film, is deposited over the sacrificial layer (FIG.2, step (1)). The sacrificial layer may be patterned to provide anchorregions 221 for the non-compliant membrane to maintain attachment to thebase substrate. As would be apparent, the anchor regions may be providedto enclose an area on the base substrate, e.g., circumscribing arectangular area with a width and a length, thereby dividing the basesubstrate into a subregion, or a window. Any suitable dimensions andnumber of windows may be provided on the base substrate.

A photoresist layer 210 may be disposed over the non-compliant membrane220 (FIG. 2, step (1)). The photoresist may be patterned into a maskthat covers select areas 212 between the anchor regions 221, therebyexposing the surface where pores are desired 213. The non-compliantmembrane may then be patterned by etching, e.g., reactive ion etching(RIE), to form the struts 222 and pores 223, e.g., micropores, throughthe non-compliant membrane in the pattern defined by the mask (FIG. 2,step (2)).

The backside of the based substrate 240 may then be patterned to removethe base substrate material over the area of the window usingfront-to-back alignment and etching, e.g., deep reactive ion etching(DRIE) (FIG. 2, step (3)). After opening the window area, thesacrificial layer 230 may be removed, e.g., using hydrofluoric acid(FIG. 2, step (4)), thereby opening the path of the pores 223 throughthe front and back sides of the non-compliant membrane 220. Theresulting non-compliant, microporous membrane 220 may be providedsupported by the base substrate 240 as a non-compliant, microporousmembrane unit 200.

With reference to FIGS. 3A-3C, processes for coating a non-compliant,porous membrane 220 with a non-porous, gas-permeable, polymeric membrane330 to fabricate a gas exchange composite membrane 300 are provided. Inan implementation of the coating process, a transitory supportingmembrane 310, e.g., a polydimethylsiloxane (PDMS) supporting membrane,is deposited on a surface of a support substrate 320, e.g., a siliconwafer, that has been surface-treated, e.g., silanized, to render thesupporting membrane detachable from the support substrate withoutsignificantly deforming the transitory supporting membrane (FIG. 3A,step (1)). For a PDMS supporting membrane, the surface of the supportsubstrate may be treated to make the surface sufficiently hydrophobic toprevent adhesion of the transitory supporting membrane to the supportsubstrate.

Then, a non-porous, gas-permeable, polymeric membrane 330, e.g., a PDMSmembrane, is deposited over the transitory supporting membrane 310 (FIG.3A, step (2)), thereby forming a two-layer, transfer membrane 332,having a top surface formed by the polymeric membrane and the bottomsurface formed by the transitory supporting membrane. Before coating thetransitory supporting membrane with the polymeric membrane, the exposedsurface of the transitory supporting membrane, i.e., the surface of thetransitory supporting membrane opposite the surface in contact with thesupport substrate 320, may be surface treated, e.g., silanized, torender the polymeric membrane detachable from the transitory supportingmembrane without significantly deforming the polymeric membrane. In thecase that a PDMS polymeric membrane is disposed on the surface of a PDMSsupporting membrane, the surface of the transitory supporting membranemay be treated, e.g., by silanization, to make the surface sufficientlyhydrophobic to prevent adhesion of the transitory supporting membrane tothe polymeric membrane. The transitory supporting membrane may bethicker than the polymeric membrane.

The transfer membrane 332 may then be detached from the supportsubstrate 320 (FIG. 3A, step (3)) using mechanical force, e.g., bypeeling. Then the top surface of the transfer membrane, i.e., theexposed surface of the polymeric membrane 330 opposite the surface thatis in contact with the transitory supporting membrane 310, is bonded tothe top surface of the non-compliant membrane 220, i.e., the surface ofthe of the membrane opposite the surface that is attached to the basesubstrate 240 of the non-compliant microporous membrane unit 200 (FIG.3A, step (4)). In order to bond the polymeric membrane to thenon-compliant membrane, the top surface of the transfer membrane and thetop surface of the non-compliant membrane may be surface-treated, e.g.,exposed to oxygen plasma, before contacting the two top surfaces witheach other. An amount of a liquid, e.g., water or isopropyl alcohol, maybe spread across the interface between the two top surfaces to promotebonding and prevent air bubble from forming between the layers.

After bonding, the polymeric membrane 330 is detached from thetransitory supporting membrane 310 (FIG. 3A, step (4)) by, e.g., peelingthe transitory supporting membrane along a direction substantiallyparallel to the top surface of the non-compliant membrane 220, therebyobtaining a composite membrane 300.

In another implementation of a process for coating a non-compliant,porous membrane 220 with a non-porous, gas-permeable, polymeric membrane330 to fabricate a composite membrane 300 (FIG. 3B), a transitorysupporting membrane 310, e.g., a polydimethylsiloxane (PDMS) supportingmembrane, is deposited on a surface of a support substrate 320, e.g., asilicon wafer, that has been surface-treated, e.g., silanized, to renderthe transitory supporting membrane detachable from the support substratewithout significantly deforming the transitory supporting membrane (FIG.3B, step (1)). For a PDMS supporting membrane, the surface of thesupport substrate may be treated, e.g., exposed to oxygen plasma, tomake the surface sufficiently hydrophilic to allow for deposition of awater-soluble polymer membrane 340.

Then, a water-soluble polymer membrane 340, e.g., a polyvinyl alcohol(PVA) membrane, is deposited over the transitory supporting membrane310, followed by a non-porous, gas-permeable, polymeric membrane 330,e.g., a PDMS membrane (FIG. 3B, step (1)), thereby forming a three-layertransfer membrane 333, having a top surface formed by the polymericmembrane, the bottom surface formed by the transitory supportingmembrane and the water-soluble polymer membrane interposed between thepolymeric and transitory supporting membranes.

The transfer membrane 333 may then be detached from the supportsubstrate 320 (FIG. 3B, step (2)) using mechanical force, e.g., bypeeling. Then the top surface of the transfer membrane, i.e., theexposed surface of the polymeric membrane 330 opposite the surface thatis in contact with the water-soluble polymer membrane 340, is bonded tothe top surface of the non-compliant membrane 220, i.e., the surface ofthe of the non-compliant membrane opposite the surface that is attachedto the base substrate 240 of the non-compliant microporous membrane unit200 (FIG. 3B, step (2)). In order to bond the polymeric membrane to thenon-compliant membrane, the top surface of the transfer membrane and thetop surface of the non-compliant membrane 220 may be surface-treated,e.g., exposed to oxygen plasma, before contacting the two top surfaceswith each other. An amount of a volatile liquid, e.g., isopropylalcohol, may be spread across the interface between the two top surfacesto promote bonding.

After bonding, the polymeric membrane 330 is detached from thetransitory supporting membrane 310 by dissolving the water-solublepolymer membrane 340 by, e.g., exposing the water-soluble polymermembrane to water, thereby obtaining a composite membrane 300 (FIG. 3B,step (3)).

Also provided herein is another process a process for coating anon-compliant, porous membrane component 200 with a non-porous,gas-permeable, polymeric membrane 330 to fabricate a composite membrane300 (FIG. 3C). Here, a transitory dissolvable film 350, e.g., a film ofSU-8 photoresist, is deposited on a surface of a support substrate 320,e.g., a silicon wafer (FIG. 3C, step (1)). The SU-8 may be spin-coatedonto the silicon wafer and the coated silicon wafer baked to form thefilm.

Following deposition of the transitory dissolvable film 350, anon-porous, gas-permeable, polymeric membrane 330, e.g., a PDMSmembrane, is deposited over the dissolvable film 350, thereby forming atwo-layer membrane on the support substrate 320 (FIG. 3C, step (2)).

Then the top surface of the two-layer membrane, i.e., the exposedsurface of the polymeric membrane 330 opposite the surface that is incontact with the transitory dissolvable film 350, is bonded to the topsurface of the non-compliant membrane 220, i.e., the surface of the ofthe non-compliant membrane opposite the surface that is attached to thebase substrate 240 of the non-compliant microporous membrane unit 200(FIG. 3C, step (3)). In order to bond the polymeric membrane to thenon-compliant membrane, the top surface of the two-layer membrane andthe top surface of the non-compliant membrane may be surface-treated,e.g., exposed to oxygen plasma, before contacting the two top surfaceswith each other. An amount of a volatile liquid, e.g., isopropylalcohol, may be spread across the interface between the two top surfacesto promote bonding.

After bonding, the polymeric membrane 330 is detached from the supportsubstrate 320 by dissolving the transitory dissolvable film 350 by,e.g., exposing the dissolvable film to a solvent, such as acetone,thereby obtaining a composite membrane 300 (FIG. 3C, step (4)).

In some embodiments, the method may further include functionalizing thefirst surface of the non-porous, gas permeable membrane, e.g., to reducefouling or coagulation of blood that comes into contact with the firstsurface, and/or to enhance diffusion of gas from the blood across themembrane. Any suitable functionality may be provided to the firstsurface using any suitable method. In some cases, the first surface isfunctionalized with polyethylene glycol (PEG), perfluorocarbon, heparinor carbonic anhydrase. Functionalizing a surface with perfluorocarbon isdescribed in, e.g., Leslie, Daniel C., et al. “A bioinspired omniphobicsurface coating on medical devices prevents thrombosis and biofouling.”Nature biotechnology (2014); functionalizing a surface with PEG isdescribed in, e.g., U.S. Pat. No. 7,695,775; U.S. App. Pub. No.20060093836; and PCT Pub. No. 2003/102133; functionalizing a surfacewith heparin is described in, e.g., PCT Pub. No. 2005/118018, all ofwhich are incorporated herein by reference.

The sacrificial layer 230 may have any suitable thickness (“a”). In somecases, the sacrificial layer has a thickness of 0.1 μm or more, e.g.,0.2 μm or more, 0.5 μm or more, 0.75 μm or more, including 1.0 μm ormore, and has a thickness of 5.0 μm or less, e.g., 4.0 μm or less, 3.0μm or less, including 2.0 μm or less. In some cases, the sacrificiallayer has a thickness in the range of 0.1 to 5.0 μm, e.g., 0.2 to 4.0 μm0.5 to 3.0 μm, including 0.75 to 2.0 μm.

The base substrate 240 may have any suitable thickness (“c”). In someembodiments, the base substrate has a thickness of 10 μm or more, e.g.,50 μm or more, 100 μm or more, 200 μm or more, 300 μm or more, including400 μm or more, and has a thickness of 1,000 μm or less, e.g., 800 μm orless, 600 μm or less, including 500 μm or less. In some cases, the basesubstrate has a thickness in the range of 10 to 1,000 μm, e.g., 50 to800 μm, 100 to 600 μm, including 200 to 600 μm.

The transitory supporting membrane 310, may have any suitable thickness(“g”). In certain embodiments, the transitory supporting membrane has athickness of 0.1 mm or more, e.g., 0.2 mm or more, 0.5 mm or more, 0.75mm or more, including 1.0 mm or more, and has a thickness of 10 mm orless, e.g., 8.0 mm or less, 6.0 mm or less, 4.0 mm or less, including3.0 mm or less. In certain embodiments, the transitory supportingmembrane has a thickness in the range of 0.1 to 10 mm, e.g., 0.2 to 8.0mm, 0.5 to 6.0 mm, including 0.75 to 6.0 mm.

The various membranes and films used in the present method may bedeposited onto a surface using any suitable method. Suitable methodsinclude, but are not limited to, spin coating, screen printing, spraycoating, solvent casting, chemical vapor deposition, and plasmadeposition. Coating a surface may further include any suitable curingand/or polymerization steps, e.g., heat curing, ultra-violet (UV)cross-linking, chemical cross-linking, etc.

Blood Oxygenation Devices and Systems

The present disclosure also provides a device for blood oxygenation,that includes a gas exchange composite membrane as described herein(FIG. 20). The blood oxygenation device 2000 may include a blood channel2010 having an inlet 2012 at a first end and an outlet 2014 at a secondend of the blood channel, as well as a gas channel 2020 having an inlet2022 at a first end and an outlet 2024 at a second end of the gaschannel. The gas exchange composite membrane 2030 may be positionedbetween the blood channel and the gas channel so as to form agas-permeable barrier between the blood channel and the gas channel,where the polymeric membrane of the composite membrane contacts theblood flowing through the blood channel, and the microporous membrane ofthe composite membrane contacts the gas flowing through the gas channel(see also FIG. 5). In general terms, deoxygenated blood entering theinlet of the blood channel can be oxygenated by oxygen diffusing fromthe gas in the gas channel through the composite membrane, as the bloodprogresses along the composite membrane toward the outlet of the bloodchannel. Carbon dioxide in the deoxygenated blood entering the inlet ofthe blood channel may also diffuse through the composite membrane fromthe blood to the gas in the gas channel, as the blood progresses alongthe composite membrane toward the outlet of the blood channel. Thus,blood exiting from the blood channel outlet may contain more oxygen andless carbon dioxide than the blood entering the blood channel at theinlet.

In some cases, the device contains two gas exchange composite membranesand two gas channels, where the gas exchange composite membranes flanktwo sides of the blood channel and a gas channel is provided for eachcomposite membrane.

The blood channel may have any suitable cross-section, where thecross-section is defined by a plane perpendicular to the averagedirection of flow of the blood flowing through the blood channel. Insome cases, the cross-section is substantially circular, oval,rectangular, square, triangular, etc. Where the cross-section isrectangular, the width of the rectangle may be any suitable length. Insome cases, the blood channel has a cross-sectional width, along theregion that overlaps with a gas diffusion window of a gas exchangecomposite membrane, of 0.5 mm or more, e.g., 1.0 mm or more, 5.0 mm ormore, 10 mm or more, 50 mm or more, including 100 mm or more, and has across-sectional width of 300 mm or less, e.g., 250 mm or less, 150 mm orless, 100 mm or less, including 90 mm or less. In some cases, the bloodchannel has a cross-sectional width, along the region that overlaps witha gas diffusion window of a gas exchange composite membrane, in therange of 0.5 to 300 mm, e.g., 1.0 to 250 mm, 5.0 to 250 mm, including5.0 to 150 mm. The height of the rectangle may be any suitable height.In some cases, the blood channel has a cross-sectional width of 0.01 mmor more, e.g., 0.05 mm or more, 0.1 mm or more, 0.15 mm or more, 0.2 mmor more, including 0.5 mm or more, and has a cross-sectional height of2.0 mm or less, e.g., 1.7 mm or less, 1.5 mm or less, 1.3 mm or less,including 1.0 mm or less. In some cases, the blood channel has across-sectional height in the range of 0.01 to 2.0 mm, e.g., 0.05 to 1.7mm, 0.1 to 1.5 mm, 0.15 to 1.3 mm, including 0.15 to 1.0 mm. The ratioof width to height of the blood channel, along the region that overlapswith a gas diffusion window of a gas exchange composite membrane, may beany suitable ratio. In some cases, the ratio of width to height of theblood channel is 10 or more, e.g., 20 or more, 50 or more, 100 or more,200 or more, including 500 or more, and is 1,000 or less, e.g., 750 orless, 600 or less, including 400 or less. In some cases, the ratio ofwidth to height of the blood channel is in the range of 10 to 1,000,e.g., 20 to 750, 50 to 600, including 100 to 400.

The length of the blood channel, from the first end to the second end ofthe blood channel, may be any suitable length. In some cases, the lengthof the blood channel is 1.0 mm or more, e.g., 5.0 mm or more, 10 mm ormore, 50 mm or more, 100 mm or more, including 200 mm or more and is 300mm or less, e.g., 280 mm or less, including 260 mm or less. In somecases, the length of the blood channel is in the range of 1.0 to 300 mm,e.g., 1.0 to 280 mm, 5.0 mm to 280 mm, including 5.0 mm to 260 mm.

The total area of the gas diffusion window of the blood oxygenationdevice may vary, and, if more than one gas exchange composite membranesare present, may be the combined area of the gas diffusion windows ofall of the gas exchange composite membranes. The total area of the gasdiffusion window of the blood oxygenation device may be 1.0 mm² or more,e.g., 10 mm² or more, 100 mm² or more, 1,000 mm² or more, 10,000 mm² ormore, including 0.1 m² or more, and may be 0.5 m² or less, e.g., 0.3 m²or less, 0.1 m² or less, 10,000 mm² or less, including 1,000 mm² orless. In some cases, the total area of the gas diffusion window of theblood oxygenation device may be in the range of 1.0 mm² to 0.5 m², e.g.,10 mm² to 0.3 m², including 100 mm² to 0.3 m².

The total volume of the blood channel may vary, and may be 1.0 mm³ ormore, e.g., 10.0 mm³ or more, 100 mm³ or more, 1,000 mm³ or more, 10,000mm³ or more, including 0.01 m³ or more, and may be 1.5 m³ or less, e.g.,1.0 m³ or less, 0.5 m³ or less, 0.3 m³ or less, including 0.1 m³ orless. In some embodiments, the volume of the blood channel may be in therange of 1 mm³ to 1.5 m³, e.g., 10 mm³ to 1.0 m³, 100 mm³ to 0.5 m³,1,000 mm³ to 0.5 m³, including 10,000 mm³ to 0.3 m³.

The average direction of the flow of blood through blood channel andaverage direction of the flow of the gas through the gas channel may bedifferent by any suitable angle. In some cases, the average direction ofthe flow of blood through blood channel and average direction of theflow of the gas through the gas channel is different by a range of 0° to10°, 10° to 20°, 20° to 30°, 30° to 40°, 40° to 50°, 50° to 60°, 60° to70°, or 80° to 90°. In some cases, the average direction of the flow ofblood through blood channel is substantially perpendicular to theaverage direction of the flow of the gas through the gas channel. Insome cases, the average direction of the flow of blood through bloodchannel is substantially parallel to the average direction of the flowof the gas through the gas channel.

In some cases, the inlet and/or outlet has the same width as the widthof the blood channel along the region that overlaps with a gas diffusionwindow of a gas exchange composite membrane. In some cases, the inletand/or outlet has a different width as the width of the blood channelalong the region that overlaps with a gas diffusion window of a gasexchange composite membrane. In some cases, the blood channel mayinclude a channel whose width tapers between an inlet and/or outlet thatis narrower than the cross-sectional width of the blood channel alongthe region that overlaps with a gas diffusion window of a gas exchangecomposite membrane.

The blood channel, e.g., the portions of the blood channel that is notformed by the gas exchange composite membrane, may be formed using anysuitable material. In some cases, the blood channel is a polymericchannel or a metal channel. In some cases, the blood channel is apolycarbonate, polyurethane or a silicone polymer channel. In somecases, the blood channel is made of PDMS. In some embodiments, the bloodchannel is made of a metal alloy, e.g., a titanium alloy.

In some cases, one or more surfaces of the blood channel arefunctionalized in any suitable manner to, e.g., reduce coagulation ofblood that comes into contact with the channel surface. In some cases, achannel surface is functionalized with polyethylene glycol (PEG).

The gas channel, e.g., the portions of the gas channel that is notformed by the gas exchange composite membrane, may be formed using anysuitable material. In some cases, the gas channel is a polymeric channelor a metal channel. In some cases, the gas channel is a polycarbonate,polyurethane, polyester or a silicone polymer channel. In some cases,the gas channel is made of PDMS or acrylic. In some embodiments, the gaschannel is made of a metal alloy, e.g., a titanium alloy.

In some embodiments, the blood oxygenation device may be a stackabledevice. Thus, in some cases, the blood oxygenation device may include ahousing with substantially flat outer surfaces to allow stacking of twoor more similar blood oxygenation devices on top of each other.

Also provided herein is a system for exchanging gas with blood, e.g.,venous blood from a subject in need of blood oxygenation support, tooxygenate the blood. The system may include an extracorporeal bloodcircuit that includes a peristaltic pump and one or more bloodoxygenation devices of the present disclosure. The system may beconfigured such that the peristaltic pump pumps blood, e.g.,deoxygenated blood, such as venous blood, from the subject to the bloodoxygenation devices and pumps back oxygenated blood into the subject'scirculatory system. Thus, the system may be a closed circuit configuredto receive venous blood, e.g., from the vena cavae, and to returnoxygenated blood to the right atrium. The system can include a gassupply unit that provides a source of an appropriate gas, e.g., a gascontaining a suitable amount of oxygen, to the blood oxygenation devicesthrough a gas conduit, e.g., tubing.

The system may further include any other suitable devices, e.g.,monitoring devices and gauges, valves, electronic control units, etc.Monitoring devices may include blood pressure gauges, blood flow meters,blood gas analyzers, such as a hemoximeter, etc. The blood gas analyzermay be an analyzer for oxygen and/or carbon dioxide. Other suitablemonitoring devices include blood monitoring devices for sulfur dioxideand/or bicarbonate content. In some cases, the present system includesmonitoring devices for the gas, including, but not limited to, gascontent analyzer (e.g., carbon monoxide sensor) or flow meters. Thesystem may also include other blood oxygenation devices that are knownin the art.

In some cases, the system includes a source of anticoagulants, e.g.,heparin. The heparin source may be any suitable heparin source. Incertain embodiments, the heparin source is a heparin pump, which may beconfigured to supply heparin to the extracorporeal circuit before theblood enters the oxygenation devices.

In certain embodiments, the components of the system are small enough tobe portable. In some cases, the blood oxygenation devices are smallenough to be wearable on the subject. Thus the blood oxygenation devicesmay be configured to be wearable, e.g. include clips, straps, hooks, orany other attachment elements to attach the device to a clothing or bodypart of the subject.

Methods of Oxygenating Blood

The present disclosure also includes a method of exchanging gas withblood, e.g., venous blood from a subject in need of blood oxygenationsupport. In general terms, the method may include pumping blood from acirculatory system of a subject to an extracorporeal blood circuit togenerate a circulating flow of the blood, e.g., flow of blood out of asuitable vein, through the extracorporeal blood circuit and back intothe subject at a suitable point in the circulatory system, e.g., intothe right atrium. The extracorporeal blood circuit includes one or morenon-circuitous blood channels of one or more blood oxygenating devices,such as the blood oxygenating devices disclosed herein. Thus, ablood-oxygenating device may include, in addition to the blood channel,a gas channel and gas exchange composite membrane(s) configured toexchange gaseous compounds between the blood and the gas across a planarsurface separating the blood channel and the gas channel. As the bloodchannel is non-circuitous, the blood can flow through the channelwithout being hindered or diverted by an obstruction in the bloodchannel, for at least 30%, e.g., at least 40%, at least 50% at least60%, at least 70%, at least 80%, at least 90%, or for substantially theentire length of the blood channel. The planar surface of the gasexchange composite membrane(s) may provide an unhindered, uniformsurface along which blood can flow and be oxygenated.

The method further includes flowing a gas containing through the gaschannel, where the gas contains any suitable amount (e.g., partialpressure) of gaseous compounds, e.g., oxygen, to provide the appropriatepressure gradient of the gaseous compound across the gas exchangecomposite membrane(s). The gas may also provide a sink to draw outgaseous compounds, e.g., carbon dioxide, dissolved in the blood.

The passage of the blood through the blood channel of the bloodoxygenating device may be controlled by the shape and dimensions of theblood channel cross-section, the shape of the path through bloodchannel, and the flow rate, such that the shear stress on the flowingblood is controlled. The shape of the blood channel cross-section, e.g.,the cross-section of the blood channel in a plane perpendicular to theaverage direction of flow of the blood, may be any suitable crosssection, and in some cases, may be rectangular. The width of therectangular cross-section, as defined by the edge of the cross-sectionthat is parallel to the gas exchange surface of the gas exchangecomposite membrane, may be any suitable width. In some cases, the bloodchannel has a cross-sectional width, along the region that overlaps withthe planar surface of the gas exchange composite membrane across whichgas exchange occurs, of 0.5 mm or more, e.g., 1.0 mm or more, 5.0 mm ormore, 10 mm or more, 50 mm or more, including 100 mm or more, and has across-sectional width of 300 mm or less, e.g., 250 mm or less, 150 mm orless, 100 mm or less, including 90 mm or less. In some cases, the bloodchannel has a cross-sectional width, along the region that overlaps withthe planar surface of the gas exchange composite membrane across whichgas exchange occurs, in the range of 0.5 to 300 mm, e.g., 1.0 to 250 mm,5.0 to 250 mm, including 5.0 to 150 mm. The height of the rectangle maybe any suitable height. In some cases, the blood channel has across-sectional width of 0.01 mm or more, e.g., 0.05 mm or more, 0.1 mmor more, 0.15 mm or more, 0.2 mm or more, including 0.5 mm or more, andhas a cross-sectional height of 2.0 mm or less, e.g., 1.7 mm or less,1.5 mm or less, 1.3 mm or less, including 1.0 mm or less. In some cases,the blood channel has a cross-sectional height in the range of 0.01 to2.0 mm, e.g., 0.05 to 1.7 mm, 0.1 to 1.5 mm, 0.15 to 1.3 mm, including0.15 to 1.0 mm. The ratio of width to height of the blood channel, alongthe region that overlaps with the planar surface of the gas exchangecomposite membrane across which gas exchange occurs, may be any suitableratio. In some cases, the ratio of width to height of the blood channelis 10 or more, e.g., 20 or more, 50 or more, 100 or more, 200 or more,including 500 or more, and is 1,000 or less, e.g., 750 or less, 600 orless, including 400 or less. In some cases, the ratio of width to heightof the blood channel is in the range of 10 to 1,000, e.g., 20 to 750, 50to 600, including 100 to 400.

Pumping the blood may include pumping the blood at a suitable flow ratethrough the extracorporeal blood circuit and/or through a bloodoxygenation device. In some cases the flow rate of the blood through theblood oxygenation device is 0.1 ml/min or more, e.g., 0.2 ml/min ormore, 0.5 ml/min or more, 1.0 ml/min or more, 2.0 ml/min or more, 5.0ml/min or more, 10 ml/min or more, 20 ml/min or more, including 50ml/min or more, and is 100 ml/min or less, 80 ml/min or less, 60 ml/minor less, 40 ml/min or less, 20 ml/min or less, including 10 ml/min orless. In some cases, the blood is pumped through the blood oxygenationdevice at a flow rate in the range of 0.1 to 100 ml/min, e.g., 0.1 to 60ml/min, 0.2 to 20 ml/min, including 0.5 to 10 ml/min.

The blood flowing through the blood channel can have any suitablemaximum shear stress. In some cases, the blood flowing through the bloodchannel has a maximum shear stress of 1,000 dyne cm⁻² or less, e.g., 800dyne cm⁻² or less, 500 dyne cm⁻² or less, 300 dyne cm⁻² or less, 250dyne cm⁻² or less, including 200 dyne cm⁻² or less.

The gaseous compound, e.g., oxygen, in the flow of gas can have anysuitable partial pressure, e.g., partial pressure relative toatmospheric pressure. In some cases, the partial pressure of the gaseouscompound in the flow of gas is 10 cmHg or more, e.g., 20 cmHg or more,50 cmHg or more, 70 cmHg or more, 90 cmHg or more, 100 cmHg or more,including 120 cmHg or more, and is 500 cmHg or less, e.g., 350 cmHg orless, 250 cmHg or less, 150 cmHg or less, including 130 cmHg or less. Insome cases, the partial pressure of the gaseous compound in the flow ofgas is in the range of 10 to 500 cmHg, e.g., 20 to 350 cmHg, 50 to 250cmHg, 90 to 250 cmHg, including 100 to 150 cmHg.

The blood oxygenating device(s) may together provide a large surfacearea for gas exchange. In some cases, the blood oxygenating device(s)collectively provide a gas exchange surface area of 0.1 m² or more,e.g., 0.2 m² or more, 0.3 m² or more, 0.5 m² or more, 0.7 m² or more,1.0 m² or more, 2.0 m² or more, 3.0 m² or more, including 4.0 m² ormore, and provide a gas exchange surface area of 5.0 m² or less, e.g.,3.5 m² or less, 2.5 m² or less, 1.5 m² or less, including 0.8 or less.In some cases, the blood oxygenating device(s) collectively provide agas exchange surface area in the range of 0.1 to 5.0 m², e.g., 0.2 to3.5 m², 0.2 to 2.5 m², 0.5 to 2.5 m², including 1.0 to 2.5 m².

The non-circuitous blood channel may provide for a suitable hydraulicpressure drop of the flow of blood before and after the bloodoxygenating device. The flow of blood across each of the non-circuitousblood channels can have a hydraulic pressure drop of 100 mmHg or less,e.g., 80 mmHg or less, 60 mmHg or less, 30 mmHg or less, 20 mmHg orless, 15 mmHg or less, 12 mmHg or less, 10 mmHg or less, including 5.0mmHg or less.

The subject can be any suitable animal subject, e.g., a mammaliansubject, such as a human, non-human primate, pig, horse, cow, sheep,dog, cat, mouse, rat, etc. In some cases, the subject is a patient inneed of blood oxygenation support, e.g., a patient with an acuterespiratory distress or a patient undergoing lung surgery. In somecases, the patient is a patient diagnosed with acute respiratorydistress syndrome (ARDS), or chronic obstructive pulmonary disease(COPD). In some cases, the patient is a patient undergoing lungtransplantation.

The present method of exchanging gas with blood can be an efficientmethod of exchanging gas with blood. In some cases, the method providesan oxygen transfer rate between the gas and the blood at each bloodoxygenation device of 0.5 mL STP/cmHg/m²/min or more, e.g., 1.0 mLSTP/cmHg/m²/min or more, 1.5 mL STP/cmHg/m²/min or more, including 2.0mL STP/cmHg/m²/min or more, and has an oxygen gas transfer rate of 10 mLSTP/cmHg/m²/min or less, e.g., 8.0 mL STP/cmHg/m²/min or less, 6.0 mLSTP/cmHg/m²/min or less, 4.0 mL STP/cmHg/m²/min or less, including 3.0mL STP/cmHg/m²/min or less, at an average blood flow rate in the rage ofabout 0.1 to about 1.0 mL/min through each blood oxygenation device. Insome embodiments, the method provides an oxygen transfer rate betweenthe gas and the blood at each blood oxygenation device in the range of0.5 to 10 mL STP/cmHg/m²/min, e.g., 1.0 to 8.0 mL STP/cmHg/m²/min, 1.5to 6.0 mL STP/cmHg/m²/min, including 1.5 to 4.0 mL STP/cmHg/m²/min, atan average blood flow rate in the rage of about 0.1 to about 1.0 mL/minthrough each blood oxygenation device.

In some cases, the method provides a carbon dioxide transfer ratebetween the gas and the blood at each blood oxygenation device of 2.0 mLSTP/cmHg/m²/min or more, e.g., 2.5 mL STP/cmHg/m²/min or more, 3.0 mLSTP/cmHg/m²/min or more, 4.0 mL STP/cmHg/m²/min or more, including 5.0mL STP/cmHg/m²/min or more, and has a carbon dioxide gas transfer rateof 50 mL STP/cmHg/m²/min or less, e.g., 40 mL STP/cmHg/m²/min or less,30 mL STP/cmHg/m²/min or less, 20 mL STP/cmHg/m²/min or less, including10 mL STP/cmHg/m²/min or less, at an average blood flow rate in the rageof about 0.1 to about 1.0 mL/min through each blood oxygenation device.In some embodiments, the method provides a carbon dioxide transfer ratebetween the gas and the blood at each blood oxygenation device in therange of 2.0 to 50 mL STP/cmHg/m²/min, e.g., 3.0 to 30 mLSTP/cmHg/m²/min, 4.0 to 20 mL STP/cmHg/m²/min, including 5.0 to 10 mLSTP/cmHg/m²/min, at an average blood flow rate in the rage of about 0.1to about 1.0 mL/min through each blood oxygenation device.

Kits

Also provided herein are kits that find use in performing methods of gasexchanging blood from a subject in need of blood oxygenation, asdescribed herein. The present kit may include a gas exchange compositemembrane of the present disclosure and a housing that includes a bloodchannel and a gas channel, wherein the housing is configured to receivethe gas exchange composite membrane between the blood channel and thegas channel, wherein the gas exchange composite membrane forms a gaspermeable barrier between the blood channel and the gas channel.

The present kit may include any other suitable component for performinga method of gas exchanging blood from a subject in need of bloodoxygenation, as described herein. In some cases, the present kitincludes instructions for using the gas exchange composite membrane inthe housing. The instructions are generally recorded on a suitablerecording medium. For example, the instructions may be printed on asubstrate, such as paper or plastic, etc. As such, the instructions maybe present in the kits as a package insert, in the labeling of thecontainer of the kit or components thereof (i.e., associated with thepackaging or subpackaging) etc. In other embodiments, the instructionsare present as an electronic storage data file present on a suitablecomputer readable storage medium, e.g. CD-ROM, digital versatile disc(DVD), flash drive, Blue-ray Disc™ etc. In yet other embodiments, theactual instructions are not present in the kit, but methods forobtaining the instructions from a remote source, e.g. via the internet,are provided. An example of this embodiment is a kit that includes a webaddress where the instructions can be viewed and/or from which theinstructions can be downloaded. As with the instructions, the methodsfor obtaining the instructions are recorded on a suitable substrate.

Components of a subject kit can be in separate containers; or can becombined in a single container.

EXAMPLES

The following examples are put forth so as to provide those of ordinaryskill in the art with a complete disclosure and description of how tomake and use the disclosed subject matter, and are not intended to limitthe scope of what the inventors regard as their invention nor are theyintended to represent that the experiments below are all or the onlyexperiments performed. Efforts have been made to ensure accuracy withrespect to numbers used (e.g. amounts, temperature, etc.) but someexperimental errors and deviations should be accounted for. Unlessindicated otherwise, parts are parts by weight, molecular weight isweight average molecular weight, temperature is in degrees Celsius, andpressure is at or near atmospheric. Standard abbreviations may be used,e.g., bp, base pair(s); kb, kilobase(s); pl, picoliter(s); s or sec,second(s); min, minute(s); h or hr, hour(s); aa, amino acid(s); kb,kilobase(s); bp, base pair(s); nt, nucleotide(s); i.m.,intramuscular(ly); i.p., intraperitoneal(ly); s.c., subcutaneous(ly);and the like.

Example 1: Silicon-Supported Extracorporeal Membrane Oxygenation in anOvine Live Animal Model

Methods

Membrane Fabrication

Silicon micropore membranes (SμMs). SμMs were produced in wafer-scalearrays of 500 nm by 4 μm rectangular slit pores. A 4-inch, 400 μm thicksilicon wafer was heated in a wet oxidation furnace for 1 hr at 1050° C.to generate a 1 μm thick thermal oxide film. The oxide layer waspatterned with anchor regions to divide the wafer into 60 μm by 240 μm“window” sub regions. A 1 μm thick polysilicon film was then depositedand patterned via photolithography and reactive ion etching (RIE) todefine the slit pores (FIG. 2, steps (1) and (2)), with a pore pitch of500 nm. The backside of the wafer was then patterned to define the samewindow dimensions using front-to-back alignment and deep reactive ionetching (DRIE) (FIG. 2, step (3)). The wafer was subsequently diced into1 cm² membranes containing 1500 windows each. Lastly, 49% hydrofluoricacid was used to remove the oxide etch-stop layer and open the slitpores (FIG. 2, step (4)).

FIG. 2. Process flow for fabrication of a silicon micropore membrane(SμM). Cross-section illustrates a single membrane “window” (not toscale). (A) Polysilicon layer is deposited and patterned on top of oxidefilm. (B) Polysilicon is etched to define 0.5 μm by 4 μm rectangularpores. (C) Backside bulk silicon is etched to open membrane window. (D)Sacrificial oxide layer is etched to open path between pores andbackside of membrane.

PDMS coating. SμMs were coated with polydimethylsiloxane (PDMS) using alift off process derived from Ingber and colleagues (Kim et al., NatureProtocols, 2013, 8:2135). First, a 4-inch silicon wafer was silanized toprevent adhesion to PDMS by exposure to trichloro(1H, 1H, 2H,2H-perfluoro-octyl)silane (Sigma-Aldrich, St. Louis, Mo., USA) vapor ina vacuum chamber for 36 hours. Sylgard® 184 PDMS (Dow Corning, Midland,Mich.) was then mixed at a monomer to crosslinker ratio of 10:1 andspin-coated onto the silicon wafer at 500 rpm for 20 seconds. The PDMSwas heat-cured at 80° C. for 2 hours to form a ˜1 mm thick film, denotedas the “thick PDMS” layer (FIG. 3A, step (1), 310). The silanizationprocess was repeated to passivate the thick PDMS.

A second PDMS layer was then prepared, using a 10:1 monomer:crosslinkerratio diluted with 50% hexanes (Sigma-Aldrich, St. Louis, Mo., USA) bymass to reduce the mixture viscosity. The PDMS mixture was spin-coatedatop the thick PDMS wafer at 6000 rpm for 5 minutes, then heat-cured at80° C. for 2 hours to form a ˜3 μm thick film, denoted as the “thinPDMS” layer (FIG. 3A, step (2), 330). After curing, the thin and thickPDMS layers were peeled from the silicon wafer as one piece (FIG. 3A,step (3)). This allowed the thick PDMS layer to act as a flexiblesupporting substrate for the thin PDMS, preventing wrinkling of the thinfilm prior to transfer to the SμM.

FIG. 3A. Process flow for polydimethylsiloxane (PDMS) bonding to SμMs.Cross-section illustrates a single membrane “window” (not to scale). (1)Thick PDMS layer (˜1 mm) is spun onto silanized silicon wafer and heatcured. (2) Thick PDMS is silanized and spin coated with hexane-dilutedPDMS to define thin PDMS film. (3) Thick and thin PDMS layers are peeledfrom silicon wafer as single sheet. (4) Thin PDMS layer is plasma-bondedto SμM; thick PDMS is peeled away. (5) Complete PDMS-SμM.

The SμM and PDMS were both exposed to oxygen plasma at 100 W for 10 s. A5 μL drop of isopropyl alcohol was deposited on the surface of the thinPDMS layer, then the SμM was settled onto the PDMS, ensuring a conformalbond as the alcohol evaporated. The PDMS and SμM were allowed to bond atroom temperature for 24 hours. The thick PDMS layer was then removed,peeling away nearly parallel to the SμM surface to minimize out-of-planestresses on the membrane (FIG. 3A, step (4)), leaving the completeasymmetric membrane unit (FIG. 3A, step (5) and FIG. 4A).

FIG. 4A. Light microscope image of PDMS-SμM.

SμM pore dimensions, membrane polysilicon thickness, and PDMS filmthickness were characterized using scanning electron microscopy (SEM).

Membrane Performance Characterization

Membrane pressure loading. Membrane mechanical robustness wasexperimentally verified by subjecting the complete PDMS-SμMs to leaktesting using a bench top hydraulic permeability testing system¹⁴, whichis capable of measuring water flux per unit pressure on the order ofnanoliters per minute in response to a transmembrane pressure gradient.Water was flowed in contact with the PDMS side of the membrane at atransmembrane pressure of 260 mmHg for 30 min. Any water that passedthrough fractures or tears in the membrane was collected and measured ina humidified chamber on a mass balance.

Gas diffusion testing. The PDMS-SμMs were tested for gas permeability ina dry bench-top flow cell connected to a pressurized gas supply and abubble flow meter (Sigma-Aldrich, St. Louis, Mo., USA), similar to asetup used by Burgess et al¹⁵. The pressurized gas, either oxygen orcarbon dioxide, was applied to one side of the membrane to form atransmembrane partial pressure gradient between 700 and 1400 mmHg. Asgas diffused across the membrane, it displaced a meniscus of detergentin the bubble flow meter, which allowed direct measurement of the volumeof gas transported across the membrane. By measuring the gas flux in thebubble flow meter, gas permeability could be calculated for thePDMS-SμMs as a function of gas flux per unit membrane area.

Blood Gas Transport

Blood channel and sweep gas channel geometry. The blood channel was3D-printed using PolyJet HD (Solid Concepts, Inc., Valencia, Calif.,USA), a urethane-based resin, to define a channel over the PDMS-SμM withdimensions of 1000 μm×800 μm×200 μm (L×W×H), tapering at the inlet andoutlet to interface with 1/16″ Luer lock tubing connectors. The sweepgas channel was machined from acrylic plastic to form a 3 mm tallchamber over the backside of the PDMS-SμM. The complete device wasassembled by compressing a 125 μm thick silicone gasket between thePDMS-SμM and the sweep gas chamber; the stack of components was securedwith machines screws. Blood flow rates through the device were chosen tomaintain wall shear stress below hemolysis thresholds.

Gas exchange ex vivo. Whole ovine blood was used in a 200 μm bloodchannel device. The blood was collected in citrate phosphate dextroseadenine (CPDA-1) bags from anesthetized sheep immediately prior toplanned termination for an unrelated experiment. Testing was conductedover a 3-hour period. Blood was maintained in a warming bath at 39° C.throughout the experiment and a peristaltic pump (Cole-Parmer, VernonHills, Ill., USA) was used to circulate blood through the device with adigital pressure gauge (General Electric Measurement & Control,Billerica, Mass., USA) monitoring blood inflow pressure continuously(FIG. 7A). The blood channel, gas channel, and PDMS-SμM were assembleddry, then primed with 20 Units/mL heparin saline for 15 minutes prior tointroduction of blood into the circuit. Pure oxygen sweep gas wassupplied to the device at a flow rate of 2.5 L/min, maintaining a sweeppressure of 10 psig. Blood flow rates were set at 0.1, 0.2, 0.5 and 1mL/min. The order of flow rates was determined using a random numbergenerator, providing for two full cycles of the four flow rates. Threeblood samples at each flow rate were collected from both the inlet andoutlet, and analyzed at six-minute intervals using an ABL5 blood gasanalyzer (Radiometer America, Westlake, Ohio, USA). The analyzerprovided pH, pCO2, pO2, SO2, HCO3 and base excess for each sample,allowing membrane exchange to be calculated using Henry's law. The bloodgas analyzer was automatically calibrated throughout the experiment.Blood was assessed for gross coagulation after each draw.

FIG. 7A. Schematic of ex vivo gas exchange experiment. Citrated bloodmaintained at 39 C in water bath was pumped through the oxygenator, withblood samples collected at the oxygenator inlet and outlet in asingle-pass manner.

Gas exchange in vivo. For in vivo testing, a similar set-up was utilized(FIG. 7B), the primary difference being the connection of the deviceinflow and outflow ports to an adult ewe with bilateral saphenous veincatheters (venous inlet right leg used 8 Fr feeding tube; venous returnleft leg used 050). The animal was sedated with a continuous drip ofketamine (0.3 mg/kg/min), diazepam (0.002 mg/kg/min), and fentanyl (1μg/kg/hr) for anesthesia and analgesia using an internal jugular veincatheter and hemodynamics were monitored continuously through a femoralarterial line; the ewe was maintained on room air for the duration ofthe experiment without advanced airway control. Animals were placed ontheir back on a heated, v-shaped table. Trained veterinary staff placedthe saphenous catheters, internal jugular catheter and administeredanesthesia. A heparin bolus of 100 Units/kg was given immediately priorto connection of the saphenous catheters to the device and a continuousinfusion of heparin at 20 Units/min from a syringe pump (KD Scientific,Holliston, Mass., USA) was used to maintain adequate anticoagulationthrough the device. At the conclusion of the experiment, the animal waseuthanized while sedated using sodium pentobarbital. The experimentprotocol was approved by the Institutional Animal Care and Use Committeeat the University of California San Francisco.

FIG. 7B. Schematic of in vivo gas exchange experiment. Venous blood froman adult ewe was withdrawn by a peristaltic pump and anti-coagulated byan inline heparin infusion pump. Blood samples were collected at theoxygenator inlet and outlet before returning to systemic circulation viaa separate venous line.

Results

Membrane Characterization

Scanning electron micrographs of the PDMS-SμMs confirmed the presence ofa conformal PDMS film with a thickness of 3.25±0.08 μm; the polysiliconsupport layer of the SμM was measured to be 0.96±0.06 μm thick (FIG.4D). SμM pore dimensions were 0.48±0.01 μm wide and 3.57±0.01 μm long.

FIG. 4D. Cross section SEM of PDMS-SμM. Blood flows in contact with thePDMS film, while sweep gas flows on the silicon membrane side, diffusingthrough the PDMS to the blood.

No water flow was detected across the PDMS-SμMs during hydraulicpermeability testing, confirming the integrity of the PDMS gas exchangemembrane.

Membrane Gas Permeability

Collection of 100 μL carbon dioxide and 10 μL oxygen gas permeate wasconducted three times for each membrane. The volumes were chosen toallow rapid collection of the target gas before the gas concentrationgradient on each side of the membrane became equalized. A set of sixPDMS-SμMs were tested, producing gas permeability results of 12.04±1.45mL STP/min/cmHg/m² for oxygen and 77.78±10.42 mL STP/min/cmHg/m² forcarbon dioxide (FIG. 8). Two of the most similar PDMS-SμMs were selectedfor comparative studies between in vivo and ex vivo blood oxygenationexperiments. For in vivo testing, the selected membrane had thefollowing permeability: 11.17±1.52 mL STP/min/cmHg/m² for O₂ and85.07±2.35 mL STP/min/cmHg/m² for CO₂. The membrane selected for ex vivotesting exhibited similar permeability: 11.19±1.81 mL STP/min/cmHg/m²for O₂ and 85.37±14.26 mL STP/min/cmHg/m² for CO₂.

FIG. 8. Membrane permeability to oxygen and carbon dioxide. Transportwas measured by diffusion from pure sample gas to room air.

Ovine Ex Vivo Testing

Citrated whole ovine blood was passed in contact with a PDMS-SμM in a200 μm high channel for 3 hours in a single-pass experiment. Bloodpressure drop across the device remained below 5 mmHg (FIG. 9) with ahomogeneous blood flow pattern for the duration of the experiment (FIG.6B). No coagulation was detected in blood drawn from the inlet or outletsample ports. The pH, pCO₂, and pO₂ of blood at the inlet sample portremained stable throughout the experiment with pH 6.97±0.12, pCO₂93.5±3.7 mmHg, and pO₂ 40.8±1.7 mmHg.

FIG. 6B. Citrated whole blood flowing through 200 μm high blood channelduring ex vivo testing.

FIG. 9. Pressure drop between oxygenator inlet and outlet(average±standard deviation) during ex vivo tests. Blood flow rates werecycled twice in random order; results displayed in chronological orderfrom left to right.

Significant oxygen exchange was seen at all flow rates except for 1mL/min (FIG. 10A). The permeability of the PDMS-SμM at each flow ratewas variable (FIG. 10B), with a general trend toward decreasing O₂permeability with time independent of flow rate. The initial observed O₂permeability was 5.97±3.13 mL STP/min/cmHg/m2. CO₂ permeability at theonset of the experiment was 19.77±12.30 mL STP/min/cmHg/m2, but noappreciable exchange was detectable thereafter (FIGS. 11A-11B).

FIGS. 10A and 10B. (FIG. 10A) Ex vivo oxygen partial pressure in bloodover time, with two cycles of randomized blood flow rates. Each bloodflow rate was maintained for 20 minutes (FIG. 10B) Ex vivo oxygenexchange rates in chronological order.

FIGS. 11A and 11B. (FIG. 11A) Ex vivo carbon dioxide partial pressure inblood. (FIG. 11B) Ex vivo carbon dioxide exchange rates in chronologicalorder.

Ovine In Vivo Testing

The pressure drop across the device varied throughout the course of thetest, ranging from 1.5 to 11.5 mmHg (FIG. 12); this variation wasindependent of flow rate, and generally increased over time. There wasno gross coagulation of blood observed throughout the experiment, butthe pattern of flow was dynamic. The animal remained hemodynamicallystable for the duration.

FIG. 12. Pressure drop between oxygenator inlet and outlet(average±standard deviation) during in vivo tests. Blood flow rates werecycled twice in random order; results displayed in chronological orderfrom left to right.

The inflow pO₂ declined over 3 hours from 54 mmHg to 37 mmHg (FIG. 13A).Oxygen permeability varied throughout the experiment with a generaltrend toward increased permeability at higher flow rates (FIG. 13B). Theoxygen permeability over time remained fairly constant throughout thecourse of the experiment with the most notable finding being increasedvariability of permeability at the 1 mL/min flow rate. The inflow pH andpCO₂ remained stable at 7.35±0.01 and 43.2±2.7 mmHg, respectively (FIG.14A). Similar to ex vivo results, there was no appreciable CO₂ exchangewhen averaged by flow rate (FIG. 14B). There was evidence of CO₂exchange during the test at a blood flow rate of 0.5 mL/min, withpermeability of 8.04±3.93 mL STP/min/cmHg/m²; CO₂ exchange wasnegligible at all subsequent blood flow rates.

FIGS. 13A and 13B. (FIG. 13A) In vivo oxygen partial pressure in bloodover time, with two cycles of randomized blood flow rates. Each bloodflow rate was maintained for 20 minutes (FIG. 13B) In vivo oxygenexchange rates in chronological order.

FIGS. 14A and 14B. (FIG. 14A) In vivo carbon dioxide partial pressure inblood. (FIG. 14B) In vivo carbon dioxide exchange rates in chronologicalorder.

Platelet and Blood Endothelial Cell Adhesion on Membranes

CD31-stained membranes showed minimal adhered cells after exposure tocitrated blood ex vivo (FIG. 15A), with less than 0.05% occlusion.However, 12.5% of membrane area was occluded by cells and plateletsafter exposure to heparinized blood in vivo (FIG. 15B).

FIGS. 15A and B. (FIG. 15A) Epifluorescence image of CD31-labeledPDMS-SμM following conclusion of ex vivo blood oxygenation experiment;<0.05% membrane surface area is occluded by adhered platelets and otherblood endothelial cells. (FIG. 15B) CD-31 labeling of adhered bloodcells on PDMS-SμM following in vivo blood oxygenation experiment showed12.5% occlusion of membrane surface area. Inset shows magnification ofthe indicated area.

Example 2: Planar Polydimethylsiloxane (PDMS)/Silicon Gas ExchangeMembranes for Respiratory Support Applications

Design

In a simplified model of gas transport across a membrane shown in Eq. 1,gas flux (Q) across a membrane with a partial pressure differencebetween the sweep gas (pO_(2,gas)) and blood sides (pO_(2,blood)) isinversely proportional to the membrane diffusive conductance, orpermeability, (S). A closer examination via Eq. 2 reveals thatpermeability is inversely proportional to membrane thickness (T). Whilematerial choice dictates the permittivity, K, for a given membrane ofarea, A, membrane thickness can be readily manipulated by adjustingfabrication parameters.

$\begin{matrix}{Q_{O_{2}} = {\left( {{pO}_{2,{gas}} - {pO}_{2,{blood}}} \right) \cdot S}} & (1) \\{S = \frac{A \cdot K}{T}} & (2)\end{matrix}$

Gas transport can therefore be enhanced by decreasing the membranethickness or increasing the partial pressure of oxygen in the sweep gasto create a steeper concentration gradient. However, a thin membranemade of highly elastic PDMS can deform due to differences between theabsolute pressures in the sweep gas and blood (P_(gas) and P_(blood),respectively), illustrated by Eq. 3, where u is deflection out of themembrane plane, E is membrane elastic modulus, and W, L, and T arewidth, length, and thickness of the membrane, respectively. This canlead to ballooning or collapse of the blood channel.

$\begin{matrix}{u \propto \frac{\left( {P_{gas} - P_{blood}} \right) \cdot W^{5}}{E \cdot T^{3} \cdot L}} & (3)\end{matrix}$

Current microfluidic platforms incorporating thin PDMS membranes usenarrow, high resistance blood channels to avoid large deformationsinduced by blood-gas pressure differences. To avoid these problems, arigid, but highly porous, silicon micropore membrane was employed as astructural support that makes a minimal contribution to the totalmembrane diffusive resistance, while allowing the use of a micron-thinPDMS film in contact with blood.

Analytical modeling was used to predict the deformation of the PDMSmembrane supported by SμMs, under a transmembrane pressure load of 77.6cmHg, or approximately 150% of the expected pressure difference betweenthe sweep gas and blood. Using the analytical solution for asimply-supported rectangular plate, Eq. 4 was used to determine themaximum deformation at the center of the PDMS membrane, u_(max), where uis the Poisson ratio of the membrane material and the coefficient0.15624 accounts for the ratio between the plate length and width. Forease of fabrication and subsequent material handling, a 4.5 μm thickPDMS film was chosen for evaluation.

$\begin{matrix}{u_{{ma}\; x} = {0.15624\frac{\left( {P_{gas} - P_{blood}} \right) \cdot \left( {1 - \upsilon^{2}} \right) \cdot W^{4}}{E \cdot T^{3}}}} & (4)\end{matrix}$

Modeling results indicated that a 4.5 μm thick PDMS film, suspended over500 nm wide by 4 μm long rectangular pores in a SμM, would deform amaximum of 0.017 nm out of plane under a 77.6 cmHg uniform pressureload. A far more significant source of deformation is expected from theSμM window component, with a calculated maximum deformation underidentical pressure loads of 351 nm. When used in an oxygenator withblood channel height on the order of 10-100 μm, this magnitude ofdeformation should have negligible effects on blood flowcharacteristics. For comparison, a 4.5 μm thick PDMS membrane spanning a1 cm channel would experience large elastic deformation and collapse thechannel under similar pressure loading.

Methods

Membrane Fabrication

Silicon micropore membranes (SμMs). SμMs were fabricated to producewafer-scale arrays of 500 nm by 4 μm rectangular slit pores. First, a 1μm thick thermally grown silicon dioxide layer was patterned with anchorregions to divide the wafer into 50 μm by 267 μm “window” sub regions. A500 nm thick polysilicon film was then deposited and patterned viaphotolithography and reactive ion etching (RIE) to define evenly spacedparallel beams (FIG. 2, steps (1) and (2)). Lines and spaces were both500 nm wide. The backside of the wafer was then patterned to define thesame window dimensions using front-to-back alignment and deep reactiveion etching (DRIE), thereby exposing the backside of the polysiliconbeams (FIG. 2, step (3)). Lastly, 49% hydrofluoric acid was used toremove the oxide etch-stop layer and open the membrane pores (FIG. 2,step (4)). For the purposes of this study, the wafers were diced to form1 cm² membranes containing 1500 windows each, with a total of 3.12e6pores per membrane. This granted an effective membrane area (die areaoccupied by windows) of 2.00e-5 m², with a window porosity of 40% andoverall die porosity of 6.24%.

PDMS coating. SμMs were coated with polydimethylsiloxane (PDMS) via aliftoff process derived from Thangawng et al. ((Weinheim an DerBergstrasse, Germany) 3, 132 (2007)) and Park et al. (Micromech Microeng19, 065016 (2009)). A 15 μm thick layer of SU-8 photoresist (MicroChemCorp., Newton, Mass.) was spin-coated onto a silicon wafer and baked(FIG. 3C, step (1)). Sylgard 184 PDMS (Dow Corning, Midland, Mich.) wasthen mixed at a monomer to crosslinker ratio of 3:1 and spin-coated ontothe photoresist (FIG. 3C, step (2)) at 4500 rpm for 2 min; the PDMS washeat-cured at 80° C. for 2 hr. The SμM was then bonded to the PDMS layerusing oxygen plasma treatment at 150 W for 5 s (FIG. 3C, step (3)); a 2μL drop of isopropyl alcohol was placed on the PDMS prior to contactwith the silicon membrane to ensure a conformal bond as the alcoholevaporated. The complete asymmetric membrane unit was released from thesilicon wafer by stripping the photoresist with acetone (FIG. 3C, step(4)).

Membrane Characterization

Membrane structure. SμM pore dimensions and membrane polysiliconthickness were measured using scanning electron microscopy (SEM). ThePDMS film thickness was measured using both SEM and a contactprofilometer (Ambios Technology, Inc., Santa Cruz, Calif.). Membranemechanical robustness was experimentally verified by subjecting thecomplete PDMS-SμMs to leak testing using a bench top hydraulicpermeability testing system, which is capable of measuring water fluxper unit pressure on the order of nanoliters per minute in response to atransmembrane pressure gradient. Water was flowed in contact with thePDMS side of the membrane at a transmembrane pressure of 260 mmHg for 30min. Membrane ruptures, or deformations greater than 1 μm out of plane,could be detected by outflow of water collected in a precision massbalance (Mettler Toledo, Columbus, Ohio).

Gas diffusion through membranes. The complete PDMS-SμMs were tested forgas permeability in a dry bench-top flow cell connected to a pressurizedgas supply and a bubble flow meter (Sigma-Aldrich, St. Louis, Mo.). Thepressurized gas, either oxygen or carbon dioxide, was applied to oneside of the membrane to form a transmembrane partial pressure gradientbetween 700 and 1400 mmHg. Gas that diffused to the opposite side of themembrane was collected in the bubble flow meter, displacing a meniscusof detergent to measure the volume of gas transported across themembrane. By measuring the gas flux per unit pressure, gas permeabilitycould be calculated for the PDMS-SμMs via Eq. 1. Membrane area used forthis calculation was the total window area (2.00e-5 m²).

Gas exchange with blood. The PDMS-SμMs were tested for gas permeabilityin a flowing blood environment using porcine whole blood, treated with 2U/mL heparin. An acrylic flow cell, containing a 1 cm² PDMS-SμM (6.24e-6m² effective gas exchange area), was used as a blood oxygenator anddeoxygenator, with the PDMS side of the membrane in direct contact withblood while the silicon backside was exposed to a sweep gas, eitheroxygen or nitrogen, respectively (FIG. 6A). A total volume of 2 mL ofblood was recirculated over the PDMS-SμM in a 50 μm high by 9 mm wide by9 mm long channel at 2 mL/min. The blood flow rate was chosen to providemaximum shear without causing hemolysis, thereby reducing the boundarylayer thickness at the membrane surface. The gas side of the membranewas first exposed to pure nitrogen at a partial pressure of 860 mmHg todeoxygenate the blood; the gas was then switched to pure oxygen tocreate a transmembrane partial pressure gradient of 1275 mmHg and driveoxygen diffusion into the blood. Oxygen partial pressure measurementswere acquired at 1 Hz using a NeoFox inline optical oxygen sensor (OceanOptics, Dunedin, Fla.), which was coated with a fluorophore coating thatwas quenched by oxygen. Total oxygen concentration was determined fromthe hemoglobin dissociation model developed by Margaria. The flow cellwas then flushed with saline solution at 2 mL/min for 5 min, thendisassembled and examined for gross clots.

FIG. 6A. Ex vivo blood oxygenation circuit with inline optical oxygensensor.

Results

Membrane Structural Characterization

SμM dimensions were characterized using scanning electron microscopy(SEM) (FIG. 4B), confirming highly uniform pore dimensions of 3.55±0.03μm long by 450±20 nm wide, with a polysilicon membrane thickness of480±20 nm. The PDMS film thickness was measured by SEM to be 4.63±0.11μm, which agreed closely with contact profilometry results.

FIG. 4B. Top view of SμM with dimensions of individual pores.

Hydraulic permeability testing of the complete PDMS-SμMs was carried outto detect membrane defects that would allow leaking of water under atransmembrane pressure gradient. Water flowed parallel to the PDMS sideof the membrane at a transmembrane pressure of 260 mmHg for 30 min, withno water flux across the membrane detected. This confirmed that therewas sufficient conformal bonding of the PDMS and SμM to prevent water orblood plasma from flowing around the PDMS and wetting the SμM pores.

Membrane Gas Permeability

Gas permeability of the PDMS-SμMs was first tested in a dry bench-topflow cell between air at ambient pressure and a pressurized pure gassource. The pressurized gas, either oxygen or carbon dioxide, wasapplied to one side of the membrane to form a transmembrane partialpressure gradient between 700 and 1400 mmHg. The volume of gas thatdiffused to the opposite side of the membrane was measured by a bubbleflow meter. Measured gas flux per unit pressure was converted to gaspermeability for the PDMS-SμMs via Eq. 1. Permeability to O₂ and CO₂were found to be 9.86±1.92 and 57.10±1.43 mL min⁻¹ m⁻² cmHg⁻¹,respectively (FIG. 16).

FIG. 16. Dry gas permeability of PDMS-SμMs.

Blood Oxygenation

Heparinized porcine whole blood was flowed over the PDMS-SμM in a 50 μmhigh by 9 mm wide by 9 mm long channel at 2 mL/min. Repeated oxygensaturation and desaturation of blood was achieved with no gross clots inthe flow cell or tubing for 3 hr, with a calculated oxygen permeabilityof 3.49±0.03 mL min⁻¹ m⁻² cmHg⁻¹ (FIG. 17). The oxygen transfer rate wasconsistent for the duration of the experiments, indicating negligiblefouling of the membrane.

FIG. 17. Comparison of membrane gas permeability in contact with dry gasand blood.

Example 3: Development of a Novel Oxygenator Membrane—a Step Toward anArtificial Lung

Methods

Membrane Design and Fabrication

Fabrication of a silicon micropore membrane (SμM). FIG. 2 illustrates asingle membrane cross-section. Polysilicon was deposited and patternedon top of silicone dioxide film (FIG. 2, step (1)). The polysilicon wasetched to define 0.5 μm by 4 μm rectangular pores (FIG. 2, step (2)).Then the backside bulk silicon was etched to open membrane window (FIG.2, step (3)). Then, the sacrificial oxide layer was etched to open pathbetween pores and back side of membrane FIG. 2, step (4).

Transfer of polydimethylsiloxane (PDMS) layer to SμMs. FIG. 3Billustrates a single membrane cross-section. A thick PDMS layer (˜1 mm),polyvinyl alcohol (10% w/w) and thin layer of PDMS (˜3 μm) were spunonto silanized silicon wafer and heat cured between each layer (FIG. 3B,step (1)). The thick PMDS-PVA-Thin PDMS layer was plasma-bonded to SμM(FIG. 3B, step (2)). The resulting construct was placed in water bathand the PVA layer dissolved (FIG. 3B, step (3)), yielding the completePDMS-SμM.

Ex-Vivo Experimental Set-Up Using Whole Blood

6 cm×1 cm SμM-PDMS membranes were placed in flow chamber to achieve gastransport in the mini-oxygenator (FIG. 5).

In a closed loop system with a peristaltic pump at a constant flow rate,initially deoxygenated blood flowed across the surface of the SμM-PDMSmembrane. A sweep gas of pure oxygen was kept at constant pressure onthe backside of the membrane. Samples of the blood were taken throughoutthe experiment and changes in oxygen concentration over time weredetermined with a bench-top hemoximeter.

Results

Oxygen Saturation in Blood

The blood oxygen saturation increased over time when exposed to theoxygenator membrane (FIG. 18).

FIG. 18. O₂ Saturation vs Time in Blood. The blood volume in the closedloop system was 18.5 ml flowing at 10 ml/min with a channel height of200 μm, and the sweep gas (O₂) pressure was kept constant around 0.53cmHg.

Determining Oxygen Permeability

The following Equation 5 was used to calculate gas permeability (k):

$\begin{matrix}{{k = \frac{\Delta\; C_{O_{2}} \times V_{blood}}{\left( {p_{O_{2,{gas}}} - p_{O_{2,{blood}}}} \right) \times t \times A_{m}}},} & (5)\end{matrix}$where C_(O2) is the concentration of O₂ (ml/l at STP), V is volume ofblood (liters), p_(O2) is the partial pressure of oxygen (cm Hg), t istime (min), A_(m) is the area of the membrane oxygenator (m²).

Average gas permeability was found to be 2.29 ml O₂ STP/min/m²/cmHg witha standard deviation of 0.37 ml O₂ STP/min/m²/cmHg (n=13) (FIG. 19).After testing membranes were inspected and found to have no visualdefects.

FIG. 19. O₂ Permeability of SμM-PDMS Membranes.

All publications and patent applications cited in this specification areherein incorporated by reference as if each individual publication orpatent application were specifically and individually indicated to beincorporated by reference. The citation of any publication is for itsdisclosure prior to the filing date and should not be construed as anadmission that the present invention is not entitled to antedate suchpublication by virtue of prior invention.

What is claimed is:
 1. A gas exchange composite membrane comprising: i)a non-porous, gas-permeable, polymeric membrane defining a first surfaceand a second surface opposite the first surface; and ii) anon-compliant, microporous membrane defining a third surface and afourth surface opposite the third surface, wherein the microporousmembrane comprises one or more gas diffusion windows, each comprising anetwork of struts defining walls of a plurality of micropores, eachmicropore extending from the third surface to the fourth surface; andthe third surface is attached to the second surface, wherein the firstsurface of the polymeric membrane provides an antithrombotic surface forgas exchange over the one or more gas diffusion windows, between bloodflowing along the first surface and a gas at the second surface.
 2. Thecomposite membrane of claim 1, wherein the first surface issubstantially flat over the one or more gas diffusion windows.
 3. Thecomposite membrane of claim 1, wherein the composite membrane has anoxygen gas permeability against air of 5 mL STP/cmHg/m²/min or more. 4.The composite membrane of claim 1, wherein the composite membrane has acarbon dioxide gas permeability against air of 20 mL STP/cmHg/m²/min ormore.
 5. The composite membrane of claim 1, wherein the compositemembrane has an oxygen gas transfer rate against blood of 0.5 mLSTP/cmHg/m²/min or more, at an average blood flow speed over the firstsurface in the range of about 1.0 to about 10 mm/sec.
 6. The compositemembrane of claim 1, wherein the composite membrane has a carbon dioxidetransfer rate against blood of 2.0 mL STP/cmHg/m²/min or more, at anaverage blood flow speed over the first surface in the range of about1.0 to about 10.0 mm/sec.
 7. The composite membrane of claim 1, whereinthe polymeric membrane has an average thickness in the range of 0.01 to100 μm.
 8. The composite membrane of claim 1, wherein the microporousmembrane has an average thickness in the range of 0.01 to 100 μm.
 9. Thecomposite membrane of claim 1, wherein each of the one or more gasdiffusion windows has a porosity in the range of 1 to 90%.
 10. Thecomposite membrane of claim 1, wherein the one or more gas diffusionwindows overlie an area in the range of 1.0 mm² to 1.0 m².
 11. Thecomposite membrane of claim 1, wherein a strut dividing adjacentmicropores of the plurality of micropores have an average width in therange of 0.005 to 10.0 μm.
 12. The composite membrane of claim 1,wherein the plurality of micropores is an array of micropores havingsubstantially uniform dimensions.
 13. The composite membrane of claim12, wherein the array is a regular array of micropores.
 14. Thecomposite membrane of claim 1, wherein a micropore of the plurality ofmicropores has an average width in the range of 0.005 to 50 μm.
 15. Thecomposite membrane of claim 14, wherein the micropore has an averagelength in the range of 0.01 to 100 μm.
 16. The composite membrane ofclaim 1, wherein the polymeric membrane is a polydimethylsiloxane(PDMS)-based polymeric membrane.
 17. The composite membrane of claim 1,wherein the first surface is functionalized with an antifouling agent,an anticoagulant and/or an enzyme.
 18. The composite membrane of claim17, wherein the first surface is functionalized with polyethyleneglycol, perfluorocarbon, heparin, polysulfobetaine methacrylate orcarbonic anhydrase.
 19. The composite membrane of claim 1, wherein themicroporous membrane is a microporous polysilicon, silicon, siliconcarbide, or silicon nitride membrane.
 20. The composite membrane ofclaim 1, wherein the fourth surface comprises an anchoring strip thatcircumscribes each gas diffusion window, wherein the anchoring stripprotrudes out relative to areas adjacent the anchoring strip on thefourth surface.
 21. The composite membrane of claim 20, wherein thecomposite membrane further comprises a base substrate attached to theanchoring strip.